text:
52 Entrées « ‹ 1 de 2 › »
Quang, Nguyen N; Goudey, S; Ségéral, E; Mohammad, A; Lemoine, S; Blugeon, C; Versapuech, M; Paillart, J C; Berlioz-Torrent, C; Emiliani, S; Gallois-Montbrun, S
Dynamic nanopore long-read sequencing analysis of HIV-1 splicing events during the early steps of infection Article de journal
Retrovirology, 17 (1), p. 25, 2020, ISBN: 32807178.
Résumé | Liens | BibTeX | Étiquettes: MARQUET PAILLART HIV RNA Alternative splicing Viral transcriptome ONT long-read sequencing, PAILLART, Unité ARN
title = {Dynamic nanopore long-read sequencing analysis of HIV-1 splicing events during the early steps of infection},
author = {N Nguyen Quang and S Goudey and E Ségéral and A Mohammad and S Lemoine and C Blugeon and M Versapuech and J C Paillart and C Berlioz-Torrent and S Emiliani and S Gallois-Montbrun},
journal = {Retrovirology},
pages = {25},
abstract = {Background
Alternative splicing is a key step in Human Immunodeficiency Virus type 1 (HIV-1) replication that is tightly regulated both temporally and spatially. More than 50 different transcripts can be generated from a single HIV-1 unspliced pre-messenger RNA (pre-mRNA) and a balanced proportion of unspliced and spliced transcripts is critical for the production of infectious virions. Understanding the mechanisms involved in the regulation of viral RNA is therefore of potential therapeutic interest. However, monitoring the regulation of alternative splicing events at a transcriptome-wide level during cell infection is challenging. Here we used the long-read cDNA sequencing developed by Oxford Nanopore Technologies (ONT) to explore in a quantitative manner the complexity of the HIV-1 transcriptome regulation in infected primary CD4+ T cells.
ONT reads mapping to the viral genome proved sufficiently long to span all possible splice junctions, even distant ones, and to be assigned to a total of 150 exon combinations. Fifty-three viral RNA isoforms, including 14 new ones were further considered for quantification. Relative levels of viral RNAs determined by ONT sequencing showed a high degree of reproducibility, compared favourably to those produced in previous reports and highly correlated with quantitative PCR (qPCR) data. To get further insights into alternative splicing regulation, we then compiled quantifications of splice site (SS) usage and transcript levels to build モsplice treesヤ, a quantitative representation of the cascade of events leading to the different viral isoforms. This approach allowed visualizing the complete rewiring of SS usages upon perturbation of SS D2 and its impact on viral isoform levels. Furthermore, we produced the first dynamic picture of the cascade of events occurring between 12 and 24 h of viral infection. In particular, our data highlighted the importance of non-coding exons in viral RNA transcriptome regulation.
ONT sequencing is a convenient and reliable strategy that enabled us to grasp the dynamic of the early splicing events modulating the viral RNA landscape in HIV-1 infected cells.
Background},
keywords = {MARQUET PAILLART HIV RNA Alternative splicing Viral transcriptome ONT long-read sequencing, PAILLART, Unité ARN},
Edelmann, F T; Schlundt, A; Heym, R G; Jenner, A; Niedner-Boblenz, A; Syed, M I; Paillart, J C; Stehle, R; Janowski, R; Sattler, M; Jansen, R P; Niessing, D
Molecular architecture and dynamics of ASH1-mRNA recognition by its mRNA-transport complex Article de journal
Nat Struct Mol Biol, 24 (2), p. 152-161, 2017, ISBN: 28092367.
Résumé | Liens | BibTeX | Étiquettes: MARQUET PAILLART Cell polarity RNA transport X-ray crystallography, PAILLART, Unité ARN
title = {Molecular architecture and dynamics of ASH1-mRNA recognition by its mRNA-transport complex},
author = {F T Edelmann and A Schlundt and R G Heym and A Jenner and A Niedner-Boblenz and M I Syed and J C Paillart and R Stehle and R Janowski and M Sattler and R P Jansen and D Niessing},
doi = {10.1038/nsmb.3351},
journal = {Nat Struct Mol Biol},
abstract = {mRNA localization is an essential mechanism of gene regulation and is required for processes such as stem-cell division, embryogenesis and neuronal plasticity. It is not known which features in the cis-acting mRNA localization elements (LEs) are specifically recognized by motor-containing transport complexes. To the best of our knowledge, no high-resolution structure is available for any LE in complex with its cognate protein complex. Using X-ray crystallography and complementary techniques, we carried out a detailed assessment of an LE of the ASH1 mRNA from yeast, its complex with its shuttling RNA-binding protein She2p, and its highly specific, cytoplasmic complex with She3p. Although the RNA alone formed a flexible stem loop, She2p binding induced marked conformational changes. However, only joining by the unstructured She3p resulted in specific RNA recognition. The notable RNA rearrangements and joint action of a globular and an unfolded RNA-binding protein offer unprecedented insights into the step-wise maturation of an mRNA-transport complex.},
keywords = {MARQUET PAILLART Cell polarity RNA transport X-ray crystallography, PAILLART, Unité ARN},
mRNA localization is an essential mechanism of gene regulation and is required for processes such as stem-cell division, embryogenesis and neuronal plasticity. It is not known which features in the cis-acting mRNA localization elements (LEs) are specifically recognized by motor-containing transport complexes. To the best of our knowledge, no high-resolution structure is available for any LE in complex with its cognate protein complex. Using X-ray crystallography and complementary techniques, we carried out a detailed assessment of an LE of the ASH1 mRNA from yeast, its complex with its shuttling RNA-binding protein She2p, and its highly specific, cytoplasmic complex with She3p. Although the RNA alone formed a flexible stem loop, She2p binding induced marked conformational changes. However, only joining by the unstructured She3p resulted in specific RNA recognition. The notable RNA rearrangements and joint action of a globular and an unfolded RNA-binding protein offer unprecedented insights into the step-wise maturation of an mRNA-transport complex.
doi:10.1038/nsmb.3351
Eckenfelder, A; Ségéral, E; Pinzon, N; Ulveling, D; Amadori, C; Charpentier, M; Nidelet, S; Concordet, J P; Zagury, J F; Paillart, J C; Berlioz-Torrent, C; Seitz, H; Emiliani, S; Gallois-Montbrun, S
Argonaute proteins regulate HIV-1 multiply spliced RNA and viral production in a Dicer independent manner. Article de journal
Nucleic Acids Res, 45 (7), p. 4158-4173, 2017, ISBN: 28003477.
Résumé | Liens | BibTeX | Étiquettes: MARQUET PAILLART, PAILLART, Unité ARN
title = {Argonaute proteins regulate HIV-1 multiply spliced RNA and viral production in a Dicer independent manner.},
author = {A Eckenfelder and E Ségéral and N Pinzon and D Ulveling and C Amadori and M Charpentier and S Nidelet and J P Concordet and J F Zagury and J C Paillart and C Berlioz-Torrent and H Seitz and S Emiliani and S Gallois-Montbrun},
doi = {10.1093/nar/gkw1289},
abstract = {Argonaute (Ago) proteins associate with microRNAs (miRNAs) to form the core of the RNA-induced silencing complex (RISC) that mediates post-transcriptional gene silencing of target mRNAs. As key players in anti-viral defense, Ago proteins are thought to have the ability to interact with human immunodeficiency virus type 1 (HIV-1) RNA. However, the role of this interaction in regulating HIV-1 replication has been debated. Here, we used high throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) to explore the interaction between Ago2 and HIV-1 RNA in infected cells. By only considering reads of 50 nucleotides length in our analysis, we identified more than 30 distinct binding sites for Ago2 along the viral RNA genome. Using reporter assays, we found four binding sites, located near splice donor sites, capable of repressing Luciferase gene expression in an Ago-dependent manner. Furthermore, inhibition of Ago1 and Ago2 levels in cells expressing HIV-1 led to an increase of viral multiply spliced transcripts and to a strong reduction in the extracellular CAp24 level. Depletion of Dicer did not affect these activities. Our results highlight a new role of Ago proteins in the control of multiply spliced HIV-1 transcript levels and viral production, independently of the miRNA pathway.},
keywords = {MARQUET PAILLART, PAILLART, Unité ARN},
Argonaute (Ago) proteins associate with microRNAs (miRNAs) to form the core of the RNA-induced silencing complex (RISC) that mediates post-transcriptional gene silencing of target mRNAs. As key players in anti-viral defense, Ago proteins are thought to have the ability to interact with human immunodeficiency virus type 1 (HIV-1) RNA. However, the role of this interaction in regulating HIV-1 replication has been debated. Here, we used high throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) to explore the interaction between Ago2 and HIV-1 RNA in infected cells. By only considering reads of 50 nucleotides length in our analysis, we identified more than 30 distinct binding sites for Ago2 along the viral RNA genome. Using reporter assays, we found four binding sites, located near splice donor sites, capable of repressing Luciferase gene expression in an Ago-dependent manner. Furthermore, inhibition of Ago1 and Ago2 levels in cells expressing HIV-1 led to an increase of viral multiply spliced transcripts and to a strong reduction in the extracellular CAp24 level. Depletion of Dicer did not affect these activities. Our results highlight a new role of Ago proteins in the control of multiply spliced HIV-1 transcript levels and viral production, independently of the miRNA pathway.
doi:10.1093/nar/gkw1289
Racine, P J; Chamontin, C; de Rocquigny, H; Bernacchi, S; Paillart, J C; Mougel, M
Requirements for nucleocapsid-mediated regulation of reverse transcription during the late steps of HIV-1 assembly. Article de journal
Sci Rep, 6 , p. 27536, 2016, ISBN: 27273064.
title = {Requirements for nucleocapsid-mediated regulation of reverse transcription during the late steps of HIV-1 assembly.},
author = {P J Racine and C Chamontin and H de Rocquigny and S Bernacchi and J C Paillart and M Mougel},
abstract = {HIV-1 is a retrovirus replicating within cells by reverse transcribing its genomic RNA (gRNA) into DNA. Within cells, virus assembly requires the structural Gag proteins with few accessory proteins, notably the viral infectivity factor (Vif) and two copies of gRNA as well as cellular factors to converge to the plasma membrane. In this process, the nucleocapsid (NC) domain of Gag binds to the packaging signal of gRNA which consists of a series of stem-loops (SL1-SL3) ensuring gRNA selection and packaging into virions. Interestingly, mutating NC activates a late-occurring reverse transcription (RT) step in producer cells, leading to the release of DNA-containing HIV-1 particles. In order to decipher the molecular mechanism regulating this late RT, we explored the role of several key partners of NC, such as Vif, gRNA and the cellular cytidine deaminase APOBEC3G that restricts HIV-1 infection by targeting the RT. By studying combinations of deletions of these putative players, we revealed that NC, SL1-SL3 and in lesser extent Vif, but not APOBEC3G, interplay regulates the late RT.},
HIV-1 is a retrovirus replicating within cells by reverse transcribing its genomic RNA (gRNA) into DNA. Within cells, virus assembly requires the structural Gag proteins with few accessory proteins, notably the viral infectivity factor (Vif) and two copies of gRNA as well as cellular factors to converge to the plasma membrane. In this process, the nucleocapsid (NC) domain of Gag binds to the packaging signal of gRNA which consists of a series of stem-loops (SL1-SL3) ensuring gRNA selection and packaging into virions. Interestingly, mutating NC activates a late-occurring reverse transcription (RT) step in producer cells, leading to the release of DNA-containing HIV-1 particles. In order to decipher the molecular mechanism regulating this late RT, we explored the role of several key partners of NC, such as Vif, gRNA and the cellular cytidine deaminase APOBEC3G that restricts HIV-1 infection by targeting the RT. By studying combinations of deletions of these putative players, we revealed that NC, SL1-SL3 and in lesser extent Vif, but not APOBEC3G, interplay regulates the late RT.
Mekdad, H E; Boutant, E; Karnib, H; Biedma, M E; Sharma, K K; Malytska, I; Laumond, G; Roy, M; Réal, E; Paillart, J C; Moog, C; Darlix, J L; Mély, Y; de Rocquigny, H
Characterization of the interaction between the HIV-1 Gag structural polyprotein and the cellular ribosomal protein L7 and its implication in viral nucleic acid remodeling. Article de journal
Résumé | Liens | BibTeX | Étiquettes: MARQUET PAILLART HIV Gag RPL7 Interaction Chaperone activity Nucleocapsid, PAILLART, Unité ARN
title = {Characterization of the interaction between the HIV-1 Gag structural polyprotein and the cellular ribosomal protein L7 and its implication in viral nucleic acid remodeling.},
author = {H E Mekdad and E Boutant and H Karnib and M E Biedma and K K Sharma and I Malytska and G Laumond and M Roy and E Réal and J C Paillart and C Moog and J L Darlix and Y Mély and H de Rocquigny},
doi = {10.1186/s12977-016-0287-4},
abstract = {Background In HIV-1 infected cells, the integrated viral DNA is transcribed by the host cell machinery to generate the full length HIV-1 RNA (FL RNA) that serves as mRNA encoding for the Gag and GagPol precursors. Virion formation is orchestrated by Gag, and the current view is that a specific interaction between newly made Gag molecules and FL RNA initiates the process. This in turn would cause FL RNA dimerization by the NC domain of Gag (GagNC). However the RNA chaperoning activity of unprocessed Gag is low as compared to the mature NC protein. This prompted us to search for GagNC co-factors. Results Here we report that RPL7, a major ribosomal protein involved in translation regulation, is a partner of Gag via its interaction with the NC domain. This interaction is mediated by the NC zinc fingers and the N- and C-termini of RPL7, respectively, but seems independent of RNA binding, Gag oligomerization and its interaction with the plasma membrane. Interestingly, RPL7 is shown for the first time to exhibit a potent DNA/RNA chaperone activity higher than that of Gag. In addition, Gag and RPL7 can function in concert to drive rapid nucleic acid hybridization. Conclusions Our results show that GagNC interacts with the ribosomal protein RPL7, favoring the notion that RPL7 could be a Gag helper chaperoning factor possibly contributing to the start of Gag assembly.},
keywords = {MARQUET PAILLART HIV Gag RPL7 Interaction Chaperone activity Nucleocapsid, PAILLART, Unité ARN},
Background In HIV-1 infected cells, the integrated viral DNA is transcribed by the host cell machinery to generate the full length HIV-1 RNA (FL RNA) that serves as mRNA encoding for the Gag and GagPol precursors. Virion formation is orchestrated by Gag, and the current view is that a specific interaction between newly made Gag molecules and FL RNA initiates the process. This in turn would cause FL RNA dimerization by the NC domain of Gag (GagNC). However the RNA chaperoning activity of unprocessed Gag is low as compared to the mature NC protein. This prompted us to search for GagNC co-factors. Results Here we report that RPL7, a major ribosomal protein involved in translation regulation, is a partner of Gag via its interaction with the NC domain. This interaction is mediated by the NC zinc fingers and the N- and C-termini of RPL7, respectively, but seems independent of RNA binding, Gag oligomerization and its interaction with the plasma membrane. Interestingly, RPL7 is shown for the first time to exhibit a potent DNA/RNA chaperone activity higher than that of Gag. In addition, Gag and RPL7 can function in concert to drive rapid nucleic acid hybridization. Conclusions Our results show that GagNC interacts with the ribosomal protein RPL7, favoring the notion that RPL7 could be a Gag helper chaperoning factor possibly contributing to the start of Gag assembly.
Valera, M S; de Rillo, Armas L; Barroso-González, J; Ziglio, S; Batisse, J; Dubois, N; Marrero-Hernández, S; Borel, S; García-Expósito, L; Biard-Piechaczyk, M; Paillart, J C; Valenzuela-Fernández, A
The HDAC6/APOBEC3G complex regulates HIV-1 infectiveness by inducing Vif autophagic degradation. Article de journal
Retrovirology, 12 , p. 53, 2015, ISBN: 26105074.
title = {The HDAC6/APOBEC3G complex regulates HIV-1 infectiveness by inducing Vif autophagic degradation.},
author = {M S Valera and L de Armas Rillo and J Barroso-González and S Ziglio and J Batisse and N Dubois and S Marrero-Hernández and S Borel and L García-Expósito and M Biard-Piechaczyk and J C Paillart and A Valenzuela-Fernández},
abstract = {BACKGROUND:
Human immunodeficiency virus type 1 (HIV-1) has evolved a complex strategy to overcome the immune barriers it encounters throughout an organism thanks to its viral infectivity factor (Vif), a key protein for HIV-1 infectivity and in vivo pathogenesis. Vif interacts with and promotes "apolipoprotein B mRNA-editing enzyme-catalytic, polypeptide-like 3G" (A3G) ubiquitination and subsequent degradation by the proteasome, thus eluding A3G restriction activity against HIV-1.
We found that cellular histone deacetylase 6 (HDAC6) directly interacts with A3G through its C-terminal BUZ domain (residues 841-1,215) to undergo a cellular co-distribution along microtubules and cytoplasm. The HDAC6/A3G complex occurs in the absence or presence of Vif, competes for Vif-mediated A3G degradation, and accounts for A3G steady-state expression level. In fact, HDAC6 directly interacts with and promotes Vif autophagic clearance, thanks to its C-terminal BUZ domain, a process requiring the deacetylase activity of HDAC6. HDAC6 degrades Vif without affecting the core binding factor β (CBF-β), a Vif-associated partner reported to be key for Vif- mediated A3G degradation. Thus HDAC6 antagonizes the proviral activity of Vif/CBF-β-associated complex by targeting Vif and stabilizing A3G. Finally, in cells producing virions, we observed a clear-cut correlation between the ability of HDAC6 to degrade Vif and to restore A3G expression, suggesting that HDAC6 controls the amount of Vif incorporated into nascent virions and the ability of HIV-1 particles of being infectious. This effect seems independent on the presence of A3G inside virions and on viral tropism.
Our study identifies for the first time a new cellular complex, HDAC6/A3G, involved in the autophagic degradation of Vif, and suggests that HDAC6 represents a new antiviral factor capable of controlling HIV-1 infectiveness by counteracting Vif and its functions.},
Our study identifies for the first time a new cellular complex, HDAC6/A3G, involved in the autophagic degradation of Vif, and suggests that HDAC6 represents a new antiviral factor capable of controlling HIV-1 infectiveness by counteracting Vif and its functions.
Borel, S; Robert-Hebmann, V; Alfaisal, J; Jain, A; Faure, M; Espert, L; Chaloin, L; Paillart, J C; Johansen, T; Biard-Piechaczyk, M
HIV-1 viral infectivity factor interacts with microtubule-associated protein light chain 3 and inhibits autophagy Article de journal
AIDS, 29 (3), p. 275-286, 2015, ISBN: 25490467.
title = {HIV-1 viral infectivity factor interacts with microtubule-associated protein light chain 3 and inhibits autophagy},
author = {S Borel and V Robert-Hebmann and J Alfaisal and A Jain and M Faure and L Espert and L Chaloin and J C Paillart and T Johansen and M Biard-Piechaczyk},
journal = {AIDS},
abstract = {Objective: Autophagy, an important anti-viral process triggered during HIV-1 entry by gp41-dependent membrane fusion, is repressed in infected CD4 T cells by an unknown mechanism. The aim of this study was to identify the role of Vif in the autophagy blockade.
Design/methods: To determine the role of Vif in autophagy inhibition, we used cell lines that express CD4 and CXCR4 and primary CD4 T cells. Pull-down experiments, immunoprecipitation assays and computational analyses were performed to analyze the interaction between Vif and LC3B, a major autophagy component, in presence or absence of the anti-viral host factor APOBEC3G, after HIV-1 infection or ectopic expression of Vif. Autophagy was analyzed after infection by viruses expressing Vif (NL4.3) or not (NL4.3ΔVif), or after exogenous Vif expression.
Results: We demonstrate that the C-terminal part of Vif interacts directly with LC3B, independently of the presence of APOBEC3G. Vif binds to pro-LC3 and Atg4-cleaved LC3 forms, and glycine 120, the amino acid conjugated to phosphatidylethanolamine on autophagosomes, is required. Importantly, we evidence that Vif inhibits autophagy during HIV-1 infection. Indeed, autophagy is detected in target cells infected by NL4.3ΔVif, but prevented in cells infected by NL4.3. Furthermore, autophagy triggered in NL4.3ΔVif-infected cells is inhibited when Vif is expressed in trans but is still active when target cells express a mutant of Vif that binds weakly to LC3B.
Conclusions: Our study unveils that Vif inhibits autophagy independently of its action on APOBEC3G and therefore suggest a new function of this viral protein in restricting innate anti-viral mechanisms.},
Objective: Autophagy, an important anti-viral process triggered during HIV-1 entry by gp41-dependent membrane fusion, is repressed in infected CD4 T cells by an unknown mechanism. The aim of this study was to identify the role of Vif in the autophagy blockade.
Conclusions: Our study unveils that Vif inhibits autophagy independently of its action on APOBEC3G and therefore suggest a new function of this viral protein in restricting innate anti-viral mechanisms.
Sleiman, D; Bernacchi, S; Guerrero, S X; Brachet, F; Larue, V; Paillart, J C; Tisne, C
Characterization of RNA binding and chaperoning activities of HIV-1 Vif protein: Importance of the C-terminal unstructured tail. Article de journal
Résumé | Liens | BibTeX | Étiquettes: MARQUET PAILLART HIV RNA chaperone Vif nucleocapsid unstructured domain, PAILLART, Unité ARN
title = {Characterization of RNA binding and chaperoning activities of HIV-1 Vif protein: Importance of the C-terminal unstructured tail.},
author = {D Sleiman and S Bernacchi and S X Guerrero and F Brachet and V Larue and J C Paillart and C Tisne},
doi = {10.4161/rna.29546},
abstract = {The viral infectivity factor (Vif) is essential for the productive infection and dissemination of HIV-1 in non-permissive cells, containing the cellular anti-HIV defense cytosine deaminases APOBEC3 (A3G and A3F). Vif neutralizes the antiviral activities of the APOBEC3G/F by diverse mechanisms including their degradation through the ubiquitin/proteasome pathway and their translational inhibition. In addition, Vif appears to be an active partner of the late steps of viral replication by interacting with Pr55Gag, reverse transcriptase and genomic RNA. Here, we expressed and purified full-length and truncated Vif proteins, and analyzed their RNA binding and chaperone properties. First, we showed by CD and NMR spectroscopies that the N-terminal domain of Vif is highly structured in solution, whereas the C-terminal domain remains mainly unfolded. Both domains exhibited substantial RNA binding capacities with dissociation constants in the nanomolar range, whereas the basic unfolded C-terminal domain of Vif was responsible in part for its RNA chaperone activity. Second, we showed by NMR chemical shift mapping that Vif and NCp7 share the same binding sites on tRNALys3, the primer of HIV-1 reverse transcriptase. Finally, our results indicate that Vif has potent RNA chaperone activity and provide direct evidence for an important role of the unstructured C-terminal domain of Vif in this capacity.},
keywords = {MARQUET PAILLART HIV RNA chaperone Vif nucleocapsid unstructured domain, PAILLART, Unité ARN},
The viral infectivity factor (Vif) is essential for the productive infection and dissemination of HIV-1 in non-permissive cells, containing the cellular anti-HIV defense cytosine deaminases APOBEC3 (A3G and A3F). Vif neutralizes the antiviral activities of the APOBEC3G/F by diverse mechanisms including their degradation through the ubiquitin/proteasome pathway and their translational inhibition. In addition, Vif appears to be an active partner of the late steps of viral replication by interacting with Pr55Gag, reverse transcriptase and genomic RNA. Here, we expressed and purified full-length and truncated Vif proteins, and analyzed their RNA binding and chaperone properties. First, we showed by CD and NMR spectroscopies that the N-terminal domain of Vif is highly structured in solution, whereas the C-terminal domain remains mainly unfolded. Both domains exhibited substantial RNA binding capacities with dissociation constants in the nanomolar range, whereas the basic unfolded C-terminal domain of Vif was responsible in part for its RNA chaperone activity. Second, we showed by NMR chemical shift mapping that Vif and NCp7 share the same binding sites on tRNALys3, the primer of HIV-1 reverse transcriptase. Finally, our results indicate that Vif has potent RNA chaperone activity and provide direct evidence for an important role of the unstructured C-terminal domain of Vif in this capacity.
doi:10.4161/rna.29546
Paillart, J C; Shehu-Xhilaga, M; Marquet, R; Mak, J
Dimerization of retroviral RNA genomes: an inseparable pair Article de journal
Nat Rev Microbiol, 2 (6), p. 461-472, 2004, ISBN: 15152202, (1740-1526 Journal Article Review Review, Tutorial).
Liens | BibTeX | Étiquettes: MARQUET, MARQUET Base Sequence Dimerization *Genome, Molecular Molecular Sequence Data Nucleic Acid Conformation Virus Replication/*genetics, PAILLART, Unité ARN, Viral HIV-1/*genetics/growth & development Human Models
title = {Dimerization of retroviral RNA genomes: an inseparable pair},
author = {J C Paillart and M Shehu-Xhilaga and R Marquet and J Mak},
journal = {Nat Rev Microbiol},
Review, Tutorial},
keywords = {MARQUET, MARQUET Base Sequence Dimerization *Genome, Molecular Molecular Sequence Data Nucleic Acid Conformation Virus Replication/*genetics, PAILLART, Unité ARN, Viral HIV-1/*genetics/growth & development Human Models},
Paillart, J C; Dettenhofer, M; Yu, X F; Ehresmann, C; Ehresmann, B; Marquet, R
First snapshots of the HIV-1 RNA structure in infected cells and in virions Article de journal
J Biol Chem, 279 (46), p. 48397-48403, 2004, ISBN: 15355993, (0021-9258 Journal Article).
title = {First snapshots of the HIV-1 RNA structure in infected cells and in virions},
author = {J C Paillart and M Dettenhofer and X F Yu and C Ehresmann and B Ehresmann and R Marquet},
pages = {48397-48403},
abstract = {With the increasing interest of RNAs in regulating a range of cell biological processes, very little is known about the structure of RNAs in tissue culture cells. We focused on the 5'-untranslated region of the human immunodeficiency virus type 1 RNA genome, a highly conserved RNA region, which contains structural domains that regulate key steps in the viral replication cycle. Up until now, structural information only came from in vitro studies. Here, we developed chemical modification assays to test nucleotide accessibility directly in infected cells and viral particles, thus circumventing possible biases and artifacts linked to in vitro assays. The secondary structure of the 5'-untranslated region in infected cells points to the existence of the various stem-loop motifs associated to distinct functions, proposed from in vitro probing, mutagenesis, and phylogeny. However, compared with in vitro data, subtle differences were observed in the dimerization initiation site hairpin, and none of the proposed long range interactions were observed between the functional domains. Moreover, no global RNA rearrangement was observed; structural differences between infected cells and viral particles were limited to the primer binding site, which became protected against chemical modification upon tRNA(3) (Lys) annealing in virions and to the main packaging signal. In addition, our data suggested that the genomic RNA could already dimerize in the cytoplasm of infected cells. Taken together, our results provided the first analysis of the dynamic of RNA structure of the human immunodeficiency virus type 1 RNA genome during virus assembly ex vivo.},
With the increasing interest of RNAs in regulating a range of cell biological processes, very little is known about the structure of RNAs in tissue culture cells. We focused on the 5'-untranslated region of the human immunodeficiency virus type 1 RNA genome, a highly conserved RNA region, which contains structural domains that regulate key steps in the viral replication cycle. Up until now, structural information only came from in vitro studies. Here, we developed chemical modification assays to test nucleotide accessibility directly in infected cells and viral particles, thus circumventing possible biases and artifacts linked to in vitro assays. The secondary structure of the 5'-untranslated region in infected cells points to the existence of the various stem-loop motifs associated to distinct functions, proposed from in vitro probing, mutagenesis, and phylogeny. However, compared with in vitro data, subtle differences were observed in the dimerization initiation site hairpin, and none of the proposed long range interactions were observed between the functional domains. Moreover, no global RNA rearrangement was observed; structural differences between infected cells and viral particles were limited to the primer binding site, which became protected against chemical modification upon tRNA(3) (Lys) annealing in virions and to the main packaging signal. In addition, our data suggested that the genomic RNA could already dimerize in the cytoplasm of infected cells. Taken together, our results provided the first analysis of the dynamic of RNA structure of the human immunodeficiency virus type 1 RNA genome during virus assembly ex vivo.
Goldschmidt, V; Paillart, J C; Rigourd, M; Ehresmann, B; Aubertin, A M; Ehresmann, C; Marquet, R
Structural variability of the initiation complex of HIV-1 reverse transcription Article de journal
title = {Structural variability of the initiation complex of HIV-1 reverse transcription},
author = {V Goldschmidt and J C Paillart and M Rigourd and B Ehresmann and A M Aubertin and C Ehresmann and R Marquet},
abstract = {HIV-1 reverse transcription is initiated from a tRNA(3)(Lys) molecule annealed to the viral RNA at the primer binding site (PBS), but the structure of the initiation complex of reverse transcription remains controversial. Here, we performed in situ structural probing, as well as in vitro structural and functional studies, of the initiation complexes formed by highly divergent isolates (MAL and NL4.3/HXB2). Our results show that the structure of the initiation complex is not conserved. In MAL, and according to sequence analysis in 14% of HIV-1 isolates, formation of the initiation complex is accompanied by complex rearrangements of the viral RNA, and extensive interactions with tRNA(3)(Lys) are required for efficient initiation of reverse transcription. In NL4.3, HXB2, and most isolates, tRNA(3)(Lys) annealing minimally affects the viral RNA structure and no interaction outside the PBS is required for optimal initiation of reverse transcription. We suggest that in MAL, extensive interactions with tRNA(3)(Lys) are required to drive the structural rearrangements generating the structural elements ultimately recognized by reverse transcriptase. In NL4.3 and HXB2, these elements are already present in the viral RNA prior to tRNA(3)(Lys) annealing, thus explaining that extensive interactions with the primer are not required. Interestingly, such interactions are required in HXB2 mutants designed to use a non-cognate tRNA as primer (tRNA(His)). In the latter case, the extended interactions are required to counteract a negative contribution associate with the alternate primer.},
HIV-1 reverse transcription is initiated from a tRNA(3)(Lys) molecule annealed to the viral RNA at the primer binding site (PBS), but the structure of the initiation complex of reverse transcription remains controversial. Here, we performed in situ structural probing, as well as in vitro structural and functional studies, of the initiation complexes formed by highly divergent isolates (MAL and NL4.3/HXB2). Our results show that the structure of the initiation complex is not conserved. In MAL, and according to sequence analysis in 14% of HIV-1 isolates, formation of the initiation complex is accompanied by complex rearrangements of the viral RNA, and extensive interactions with tRNA(3)(Lys) are required for efficient initiation of reverse transcription. In NL4.3, HXB2, and most isolates, tRNA(3)(Lys) annealing minimally affects the viral RNA structure and no interaction outside the PBS is required for optimal initiation of reverse transcription. We suggest that in MAL, extensive interactions with tRNA(3)(Lys) are required to drive the structural rearrangements generating the structural elements ultimately recognized by reverse transcriptase. In NL4.3 and HXB2, these elements are already present in the viral RNA prior to tRNA(3)(Lys) annealing, thus explaining that extensive interactions with the primer are not required. Interestingly, such interactions are required in HXB2 mutants designed to use a non-cognate tRNA as primer (tRNA(His)). In the latter case, the extended interactions are required to counteract a negative contribution associate with the alternate primer.
Ennifar, E; Paillart, J C; Marquet, R; Ehresmann, B; Ehresmann, C; Dumas, P; Walter, P
HIV-1 RNA dimerization initiation site is structurally similar to the ribosomal A site and binds aminoglycoside antibiotics Article de journal
J Biol Chem, 278 (4), p. 2723-2730, 2003, ISBN: 12435744, (0021-9258 Journal Article).
Résumé | Liens | BibTeX | Étiquettes: Anti-Bacterial Agents/*pharmacology Binding Sites Dimerization HIV-1/*metabolism Models, ENNIFAR, MARQUET, Molecular Neomycin/pharmacology Nucleic Acid Conformation Paromomycin/pharmacology Protein Binding RNA/metabolism *RNA, Non-U.S. Gov't Temperature Ultraviolet Rays, PAILLART, Unité ARN, Viral Ribosomes/*metabolism Support
title = {HIV-1 RNA dimerization initiation site is structurally similar to the ribosomal A site and binds aminoglycoside antibiotics},
author = {E Ennifar and J C Paillart and R Marquet and B Ehresmann and C Ehresmann and P Dumas and P Walter},
abstract = {Human immunodeficiency virus (HIV) genomic RNA is packaged into virions as a dimer. The first step of dimerization is the formation of a kissing-loop complex at the so-called dimerization initiation site (DIS). We found an unexpected and fortuitous resemblance between the HIV-1 DIS kissing-loop complex and the eubacterial 16 S ribosomal aminoacyl-tRNA site (A site), which is the target of aminoglycoside antibiotics. Similarities exist not only at the primary and secondary structure level but also at the tertiary structure level, as revealed by comparison of the respective DIS and A site crystal structures. Gel shift, inhibition of lead-induced cleavage, and footprinting experiments showed that paromomycin and neomycin specifically bind to the kissing-loop complex formed by the DIS, with an affinity and a geometry similar to that observed for the A site. Modeling of the aminoglycoside-DIS complex allowed us to identify antibiotic modifications likely to increase the affinity and/or the specificity for the DIS. This could be a starting point for designing antiviral drugs against HIV-1 RNA dimerization.},
keywords = {Anti-Bacterial Agents/*pharmacology Binding Sites Dimerization HIV-1/*metabolism Models, ENNIFAR, MARQUET, Molecular Neomycin/pharmacology Nucleic Acid Conformation Paromomycin/pharmacology Protein Binding RNA/metabolism *RNA, Non-U.S. Gov't Temperature Ultraviolet Rays, PAILLART, Unité ARN, Viral Ribosomes/*metabolism Support},
Human immunodeficiency virus (HIV) genomic RNA is packaged into virions as a dimer. The first step of dimerization is the formation of a kissing-loop complex at the so-called dimerization initiation site (DIS). We found an unexpected and fortuitous resemblance between the HIV-1 DIS kissing-loop complex and the eubacterial 16 S ribosomal aminoacyl-tRNA site (A site), which is the target of aminoglycoside antibiotics. Similarities exist not only at the primary and secondary structure level but also at the tertiary structure level, as revealed by comparison of the respective DIS and A site crystal structures. Gel shift, inhibition of lead-induced cleavage, and footprinting experiments showed that paromomycin and neomycin specifically bind to the kissing-loop complex formed by the DIS, with an affinity and a geometry similar to that observed for the A site. Modeling of the aminoglycoside-DIS complex allowed us to identify antibiotic modifications likely to increase the affinity and/or the specificity for the DIS. This could be a starting point for designing antiviral drugs against HIV-1 RNA dimerization.
Paillart, J C; Skripkin, E; Ehresmann, B; Ehresmann, C; Marquet, R
In vitro evidence for a long range pseudoknot in the 5'-untranslated and matrix coding regions of HIV-1 genomic RNA Article de journal
Résumé | Liens | BibTeX | Étiquettes: 5' Untranslated Regions/*genetics Base Sequence Dimerization HIV-1/*genetics Human Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Oligodeoxyribonucleotides, Antisense/pharmacology Poly A/chemistry/genetics RNA, MARQUET, Non-U.S. Gov't Virus Replication/genetics, Nucleic Acid Support, PAILLART, Unité ARN, Viral/chemistry/drug effects/*genetics Sequence Alignment Sequence Homology
title = {In vitro evidence for a long range pseudoknot in the 5'-untranslated and matrix coding regions of HIV-1 genomic RNA},
author = {J C Paillart and E Skripkin and B Ehresmann and C Ehresmann and R Marquet},
abstract = {The 5'-untranslated leader region of human immunodeficiency virus type 1 (HIV-1) RNA contains multiple signals that control distinct steps of the viral replication cycle such as transcription, reverse transcription, genomic RNA dimerization, splicing, and packaging. It is likely that fine tuned coordinated regulation of these functions is achieved through specific RNA-protein and RNA-RNA interactions. In a search for cis-acting elements important for the tertiary structure of the 5'-untranslated region of HIV-1 genomic RNA, we identified, by ladder selection experiments, a short stretch of nucleotides directly downstream of the poly(A) signal that interacts with a nucleotide sequence located in the matrix region. Confirmation of the sequence of the interacting sites was obtained by partial or complete inhibition of this interaction by antisense oligonucleotides and by nucleotide substitutions. In the wild type RNA, this long range interaction was intramolecular, since no intermolecular RNA association was detected by gel electrophoresis with an RNA mutated in the dimerization initiation site and containing both sequences involved in the tertiary interaction. Moreover, the functional importance of this interaction is supported by its conservation in all HIV-1 isolates as well as in HIV-2 and simian immunodeficiency virus. Our results raise the possibility that this long range RNA-RNA interaction might be involved in the full-length genomic RNA selection during packaging, repression of the 5' polyadenylation signal, and/or splicing regulation.},
keywords = {5' Untranslated Regions/*genetics Base Sequence Dimerization HIV-1/*genetics Human Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Oligodeoxyribonucleotides, Antisense/pharmacology Poly A/chemistry/genetics RNA, MARQUET, Non-U.S. Gov't Virus Replication/genetics, Nucleic Acid Support, PAILLART, Unité ARN, Viral/chemistry/drug effects/*genetics Sequence Alignment Sequence Homology},
The 5'-untranslated leader region of human immunodeficiency virus type 1 (HIV-1) RNA contains multiple signals that control distinct steps of the viral replication cycle such as transcription, reverse transcription, genomic RNA dimerization, splicing, and packaging. It is likely that fine tuned coordinated regulation of these functions is achieved through specific RNA-protein and RNA-RNA interactions. In a search for cis-acting elements important for the tertiary structure of the 5'-untranslated region of HIV-1 genomic RNA, we identified, by ladder selection experiments, a short stretch of nucleotides directly downstream of the poly(A) signal that interacts with a nucleotide sequence located in the matrix region. Confirmation of the sequence of the interacting sites was obtained by partial or complete inhibition of this interaction by antisense oligonucleotides and by nucleotide substitutions. In the wild type RNA, this long range interaction was intramolecular, since no intermolecular RNA association was detected by gel electrophoresis with an RNA mutated in the dimerization initiation site and containing both sequences involved in the tertiary interaction. Moreover, the functional importance of this interaction is supported by its conservation in all HIV-1 isolates as well as in HIV-2 and simian immunodeficiency virus. Our results raise the possibility that this long range RNA-RNA interaction might be involved in the full-length genomic RNA selection during packaging, repression of the 5' polyadenylation signal, and/or splicing regulation.
Paillart, J C; Westhof, E; Ehresmann, C; Ehresmann, B; Marquet, R
Non-canonical interactions in a kissing loop complex: the dimerization initiation site of HIV-1 genomic RNA Article de journal
J Mol Biol, 270 (1), p. 36-49, 1997, ISBN: 9231899, (0022-2836 Journal Article).
Résumé | Liens | BibTeX | Étiquettes: Dimerization HIV-1/*genetics Kinetics Models, MARQUET, Molecular Mutagenesis, Non-U.S. Gov't, PAILLART, Site-Directed Nucleic Acid Conformation Purines/chemistry RNA, Unité ARN, Viral/*chemistry/genetics/*metabolism Support
title = {Non-canonical interactions in a kissing loop complex: the dimerization initiation site of HIV-1 genomic RNA},
author = {J C Paillart and E Westhof and C Ehresmann and B Ehresmann and R Marquet},
url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9231899},
isbn = {9231899},
abstract = {Retroviruses encapsidate two molecules of genomic RNA that are noncovalently linked close to their 5' ends in a region called the dimer linkage structure (DLS). The dimerization initiation site (DIS) of human immunodeficiency virus type 1 (HIV-1) constitutes the essential part of the DLS in vitro and is crucial for efficient HIV-1 replication in cell culture. We previously identified the DIS as a hairpin structure, located upstream of the major splice donor site, that contains in the loop a six-nucleotide self-complementary sequence preceded and followed by two and one purines, respectively. Two RNA monomers form a kissing loop complex via intermolecular interactions of the six nucleotide self-complementary sequence. Here, we introduced compensatory mutations in the self-complementary sequence and/or a mutation in the flanking purines. We determined the kinetics of dimerization, the thermal stabilities and the apparent equilibrium dissociation constants of wild-type and mutant dimers and used chemical probing to obtain structural information. Our results demonstrate the importance of the 5'-flanking purine and of the two central bases of the self-complementary sequence in the dimerization process. The experimental data are rationalized by triple interactions between these residues in the deep groove of the kissing helix and are incorporated into a three-dimensional model of the kissing loop dimer. In addition, chemical probing and molecular modeling favor the existence of a non-canonical interaction between the conserved adenine residues at the first and last positions in the DIS loop. Furthermore, we show that destabilization of the kissing loop complex at the DIS can be compensated by interactions involving sequences located downstream of the splice donor site of the HIV-1 genomic RNA.},
keywords = {Dimerization HIV-1/*genetics Kinetics Models, MARQUET, Molecular Mutagenesis, Non-U.S. Gov't, PAILLART, Site-Directed Nucleic Acid Conformation Purines/chemistry RNA, Unité ARN, Viral/*chemistry/genetics/*metabolism Support},
Retroviruses encapsidate two molecules of genomic RNA that are noncovalently linked close to their 5' ends in a region called the dimer linkage structure (DLS). The dimerization initiation site (DIS) of human immunodeficiency virus type 1 (HIV-1) constitutes the essential part of the DLS in vitro and is crucial for efficient HIV-1 replication in cell culture. We previously identified the DIS as a hairpin structure, located upstream of the major splice donor site, that contains in the loop a six-nucleotide self-complementary sequence preceded and followed by two and one purines, respectively. Two RNA monomers form a kissing loop complex via intermolecular interactions of the six nucleotide self-complementary sequence. Here, we introduced compensatory mutations in the self-complementary sequence and/or a mutation in the flanking purines. We determined the kinetics of dimerization, the thermal stabilities and the apparent equilibrium dissociation constants of wild-type and mutant dimers and used chemical probing to obtain structural information. Our results demonstrate the importance of the 5'-flanking purine and of the two central bases of the self-complementary sequence in the dimerization process. The experimental data are rationalized by triple interactions between these residues in the deep groove of the kissing helix and are incorporated into a three-dimensional model of the kissing loop dimer. In addition, chemical probing and molecular modeling favor the existence of a non-canonical interaction between the conserved adenine residues at the first and last positions in the DIS loop. Furthermore, we show that destabilization of the kissing loop complex at the DIS can be compensated by interactions involving sequences located downstream of the splice donor site of the HIV-1 genomic RNA.
Skripkin, E; Paillart, J C; Marquet, R; Blumenfeld, M; Ehresmann, B; Ehresmann, C
Mechanisms of inhibition of in vitro dimerization of HIV type I RNA by sense and antisense oligonucleotides Article de journal
J Biol Chem, 271 (46), p. 28812-28817, 1996, ISBN: 8910525, (0021-9258 Journal Article).
Résumé | Liens | BibTeX | Étiquettes: Antisense/*pharmacology RNA, Biopolymers HIV-1/*genetics Nucleic Acid Conformation Oligonucleotides/*pharmacology Oligonucleotides, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*antagonists & inhibitors/chemistry Support
title = {Mechanisms of inhibition of in vitro dimerization of HIV type I RNA by sense and antisense oligonucleotides},
author = {E Skripkin and J C Paillart and R Marquet and M Blumenfeld and B Ehresmann and C Ehresmann},
abstract = {Retroviruses display a strong selective pressure to maintain the dimeric nature of their genomic RNAs, suggesting that dimerization is essential for viral replication. Recently, we identified the cis-element required for initiation of human immunodeficiency virus type I (HIV-I) RNA dimerization in vitro. The dimerization initiation site (DIS) is a hairpin structure containing a self-complementary sequence in the loop. We proposed that dimerization is initiated by a loop-loop kissing interaction involving the self-complementary sequence present in each monomer. We tested the ability of sense and antisense oligonucleotides targeted against the DIS to interfere with a preformed viral RNA dimer. Self-dimerization and inhibition properties of the tested oligonucleotides are dictated by the nature of the loop. An RNA loop is absolutely required in the case of sense oligonucleotides, whereas the nature and the sequence of the stem is not important. They form reversible loop-loop interactions and act as competitive inhibitors. Antisense oligonucleotides are less efficient in self-dimerization and are more potent inhibitors than sense oligonucleotides. They are less sensitive to the nature of the loop than the antisense oligonucleotides. Antisense hairpins with either RNA or DNA stems are able to form highly stable and irreversible complexes with viral RNA, resulting from complete extension of base pairing initiated by loop-loop interaction.},
keywords = {Antisense/*pharmacology RNA, Biopolymers HIV-1/*genetics Nucleic Acid Conformation Oligonucleotides/*pharmacology Oligonucleotides, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*antagonists & inhibitors/chemistry Support},
Retroviruses display a strong selective pressure to maintain the dimeric nature of their genomic RNAs, suggesting that dimerization is essential for viral replication. Recently, we identified the cis-element required for initiation of human immunodeficiency virus type I (HIV-I) RNA dimerization in vitro. The dimerization initiation site (DIS) is a hairpin structure containing a self-complementary sequence in the loop. We proposed that dimerization is initiated by a loop-loop kissing interaction involving the self-complementary sequence present in each monomer. We tested the ability of sense and antisense oligonucleotides targeted against the DIS to interfere with a preformed viral RNA dimer. Self-dimerization and inhibition properties of the tested oligonucleotides are dictated by the nature of the loop. An RNA loop is absolutely required in the case of sense oligonucleotides, whereas the nature and the sequence of the stem is not important. They form reversible loop-loop interactions and act as competitive inhibitors. Antisense oligonucleotides are less efficient in self-dimerization and are more potent inhibitors than sense oligonucleotides. They are less sensitive to the nature of the loop than the antisense oligonucleotides. Antisense hairpins with either RNA or DNA stems are able to form highly stable and irreversible complexes with viral RNA, resulting from complete extension of base pairing initiated by loop-loop interaction.
A loop-loop "kissing" complex is the essential part of the dimer linkage of genomic HIV-1 RNA Article de journal
Proc Natl Acad Sci U S A, 93 (11), p. 5572-5577, 1996, ISBN: 8643617, (0027-8424 Journal Article).
Résumé | Liens | BibTeX | Étiquettes: Base Composition Base Sequence HIV-1/*genetics Heat Human Kinetics Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Nucleic Acid Denaturation Plasmids RNA, Genetic, MARQUET, Non-U.S. Gov't Thermodynamics Transcription, PAILLART, Unité ARN, Viral/biosynthesis/*chemistry/*metabolism Support
title = {A loop-loop "kissing" complex is the essential part of the dimer linkage of genomic HIV-1 RNA},
journal = {Proc Natl Acad Sci U S A},
abstract = {RNA-RNA interactions govern a number of biological processes. Several RNAs, including natural sense and antisense RNAs, interact by means of a two-step mechanism: recognition is mediated by a loop-loop complex, which is then stabilized by formation of an extended intermolecular duplex. It was proposed that the same mechanism holds for dimerization of the genomic RNA of human immunodeficiency virus type 1 (HIV-1), an event thought to control crucial steps of HIV-1 replication. However, whereas interaction between the partially self-complementary loop of the dimerization initiation site (DIS) of each monomer is well established, formation of the extended duplex remained speculative. Here we first show that in vitro dimerization of HIV-1 RNA is a specific process, not resulting from simple annealing of denatured molecules. Next we used mutants of the DIS to test the formation of the extended duplex. Four pairs of transcomplementary mutants were designed in such a way that all pairs can form the loop-loop "kissing" complex, but only two of them can potentially form the extended duplex. All pairs of mutants form heterodimers whose thermal stability, dissociation constant, and dynamics were analyzed. Taken together, our results indicate that, in contrast with the interactions between natural sense and antisense RNAs, no extended duplex is formed during dimerization of HIV-1 RNA. We also showed that 55-mer sense RNAs containing the DIS are able to interfere with the preformed HIV-1 RNA dimer.},
keywords = {Base Composition Base Sequence HIV-1/*genetics Heat Human Kinetics Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Nucleic Acid Denaturation Plasmids RNA, Genetic, MARQUET, Non-U.S. Gov't Thermodynamics Transcription, PAILLART, Unité ARN, Viral/biosynthesis/*chemistry/*metabolism Support},
RNA-RNA interactions govern a number of biological processes. Several RNAs, including natural sense and antisense RNAs, interact by means of a two-step mechanism: recognition is mediated by a loop-loop complex, which is then stabilized by formation of an extended intermolecular duplex. It was proposed that the same mechanism holds for dimerization of the genomic RNA of human immunodeficiency virus type 1 (HIV-1), an event thought to control crucial steps of HIV-1 replication. However, whereas interaction between the partially self-complementary loop of the dimerization initiation site (DIS) of each monomer is well established, formation of the extended duplex remained speculative. Here we first show that in vitro dimerization of HIV-1 RNA is a specific process, not resulting from simple annealing of denatured molecules. Next we used mutants of the DIS to test the formation of the extended duplex. Four pairs of transcomplementary mutants were designed in such a way that all pairs can form the loop-loop "kissing" complex, but only two of them can potentially form the extended duplex. All pairs of mutants form heterodimers whose thermal stability, dissociation constant, and dynamics were analyzed. Taken together, our results indicate that, in contrast with the interactions between natural sense and antisense RNAs, no extended duplex is formed during dimerization of HIV-1 RNA. We also showed that 55-mer sense RNAs containing the DIS are able to interfere with the preformed HIV-1 RNA dimer.
The use of chemical modification interference and inverse PCR mutagenesis to identify the dimerization initiation site of HIV-1 genomic RNA Article de journal
Pharm Acta Helv, 71 (1), p. 21-28, 1996, ISBN: 8786995, (0031-6865 Journal Article).
Résumé | Liens | BibTeX | Étiquettes: Base Sequence *Genome, MARQUET, Non-U.S. Gov't, PAILLART, Site-Directed Polymerase Chain Reaction RNA, Unité ARN, Viral HIV-1/*chemistry Human Molecular Sequence Data Mutagenesis, Viral/*chemistry Support
title = {The use of chemical modification interference and inverse PCR mutagenesis to identify the dimerization initiation site of HIV-1 genomic RNA},
journal = {Pharm Acta Helv},
abstract = {The retroviral genome consists of two identical RNA molecules physically linked together close to their 5' end, in a region called the Dimer Linkage Structure (DLS). Recent findings suggest that dimerization is involved in encapsidation, regulation of translation and reverse transcription. Previous in vitro studies localized the DLS of HIV-1 in a region downstream of the splice donor (SD) site. More recently, we showed that dimerization of HIV-1 RNA also involves sequences upstream of the SD site. Modification interference experiments and site-directed mutagenesis were used to identify the nucleotides required in the dimerization process of HIV-1 RNA. Our results point out a self-complementary sequence located in a hairpin loop, between the Primer Binding Site (PBS) and the SD site, as the Dimerization Initiation Site.},
keywords = {Base Sequence *Genome, MARQUET, Non-U.S. Gov't, PAILLART, Site-Directed Polymerase Chain Reaction RNA, Unité ARN, Viral HIV-1/*chemistry Human Molecular Sequence Data Mutagenesis, Viral/*chemistry Support},
The retroviral genome consists of two identical RNA molecules physically linked together close to their 5' end, in a region called the Dimer Linkage Structure (DLS). Recent findings suggest that dimerization is involved in encapsidation, regulation of translation and reverse transcription. Previous in vitro studies localized the DLS of HIV-1 in a region downstream of the splice donor (SD) site. More recently, we showed that dimerization of HIV-1 RNA also involves sequences upstream of the SD site. Modification interference experiments and site-directed mutagenesis were used to identify the nucleotides required in the dimerization process of HIV-1 RNA. Our results point out a self-complementary sequence located in a hairpin loop, between the Primer Binding Site (PBS) and the SD site, as the Dimerization Initiation Site.
Paillart, J C; Marquet, R; Skripkin, E; Ehresmann, C; Ehresmann, B
Dimerization of retroviral genomic RNAs: structural and functional implications Article de journal
Biochimie, 78 (7), p. 639-653, 1996, ISBN: 8955907, (0300-9084 Journal Article Review Review, Academic).
Résumé | Liens | BibTeX | Étiquettes: Animals Base Sequence HIV-1/genetics Human Microscopy, DNA Support, Electron Molecular Sequence Data *Nucleic Acid Conformation RNA, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*chemistry/metabolism Rats Retroviridae/*genetics Sequence Analysis
title = {Dimerization of retroviral genomic RNAs: structural and functional implications},
author = {J C Paillart and R Marquet and E Skripkin and C Ehresmann and B Ehresmann},
abstract = {Retroviruses are a family of widespread small animal viruses at the origin of a diversity of diseases. They share common structural and functional properties such as reverse transcription of their RNA genome and integration of the proviral DNA into the host genome, and have the particularity of packaging a diploid genome. The genome of all retroviruses is composed of two homologous RNA molecules that are non-covalently linked near their 5' end in a region called the dimer linkage structure (DLS). There is now considerable evidence that a specific site (or sites) in the 5' leader region of all retroviruses, located either upstream or/and downstream of the major splice donor site, is involved in the dimer linkage. For MoMuLV and especially HIV-1, it was shown that dimerization is initiated at a stem-loop structure named the dimerization initiation site (DIS). The DIS of HIV-1 and related regions in other retroviruses corresponds to a highly conserved structure with a self-complementary loop sequence, that is involved in a typical loop-loop 'kissing' complex which can be further stabilized by long distance interactions or by conformational rearrangements. RNA interactions involved in the viral RNA dimer were postulated to regulate several key steps in retroviral cycle, such as: i) translation and encapsidation: the arrest of gag translation imposed by the highly structured DLS-encapsidation signal would leave the RNA genome available for the encapsidation machinery; and ii) recombination during reverse transcription: the presence of two RNA molecules in particles would be necessary for variability and viability of virus progeny and the ordered structure imposed by the DLS would be required for efficient reverse transcription.},
Review, Academic},
keywords = {Animals Base Sequence HIV-1/genetics Human Microscopy, DNA Support, Electron Molecular Sequence Data *Nucleic Acid Conformation RNA, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*chemistry/metabolism Rats Retroviridae/*genetics Sequence Analysis},
Retroviruses are a family of widespread small animal viruses at the origin of a diversity of diseases. They share common structural and functional properties such as reverse transcription of their RNA genome and integration of the proviral DNA into the host genome, and have the particularity of packaging a diploid genome. The genome of all retroviruses is composed of two homologous RNA molecules that are non-covalently linked near their 5' end in a region called the dimer linkage structure (DLS). There is now considerable evidence that a specific site (or sites) in the 5' leader region of all retroviruses, located either upstream or/and downstream of the major splice donor site, is involved in the dimer linkage. For MoMuLV and especially HIV-1, it was shown that dimerization is initiated at a stem-loop structure named the dimerization initiation site (DIS). The DIS of HIV-1 and related regions in other retroviruses corresponds to a highly conserved structure with a self-complementary loop sequence, that is involved in a typical loop-loop 'kissing' complex which can be further stabilized by long distance interactions or by conformational rearrangements. RNA interactions involved in the viral RNA dimer were postulated to regulate several key steps in retroviral cycle, such as: i) translation and encapsidation: the arrest of gag translation imposed by the highly structured DLS-encapsidation signal would leave the RNA genome available for the encapsidation machinery; and ii) recombination during reverse transcription: the presence of two RNA molecules in particles would be necessary for variability and viability of virus progeny and the ordered structure imposed by the DLS would be required for efficient reverse transcription.
Paillart, J C; Berthoux, L; Ottmann, M; Darlix, J L; Marquet, R; Ehresmann, B; Ehresmann, C
A dual role of the putative RNA dimerization initiation site of human immunodeficiency virus type 1 in genomic RNA packaging and proviral DNA synthesis Article de journal
J Virol, 70 (12), p. 8348-8354, 1996, ISBN: 8970954, (0022-538x Journal Article).
Résumé | Liens | BibTeX | Étiquettes: Animals COS Cells DNA, Genetic Virion *Virus Assembly, MARQUET, Non-U.S. Gov't Transcription, Nucleic Acid Support, PAILLART, Post-Translational Proviruses/genetics *RNA, Unité ARN, Viral *Regulatory Sequences, Viral HIV-1/*genetics/physiology Human Mutagenesis Protein Processing, Viral/*biosynthesis Gene Expression Genome
title = {A dual role of the putative RNA dimerization initiation site of human immunodeficiency virus type 1 in genomic RNA packaging and proviral DNA synthesis},
author = {J C Paillart and L Berthoux and M Ottmann and J L Darlix and R Marquet and B Ehresmann and C Ehresmann},
journal = {J Virol},
abstract = {In retroviruses, the genomic RNA is in the form of a 60S-70S complex composed of two identical genome-length RNA molecules tightly associated through numerous interactions. A major interaction, called the dimer linkage structure, has been found near the RNA 5' end and is probably involved in the control of translation, packaging, and recombination during proviral DNA synthesis. Recently, a small sequence corresponding to a stem-loop structure located in the 5' leader of human immunodeficiency virus type 1 (HIV-1) RNA was found to be required for the initiation of HIV-1 RNA dimerization in vitro and named the dimerization initiation site (E. Skripkin, J.-C. Paillart, R. Marquet, B. Ehresmann, and C. Ehresmann, Proc. Natl. Acad. Sci. USA 91: 4945-4949, 1994). To investigate the possible role of this 5' stem-loop in HIV-1 virion formation and infectivity, four mutant viruses were generated and analyzed in vivo. Results show that deletion of the stem-loop structure reduces infectivity by a factor of 10(3) whereas loop substitutions cause a decrease of 10- to 100-fold. The level of genomic RNA packaging was found to be decreased fivefold in mutants virions containing the stem-loop deletion and only twofold in the loop-substituted virions. Surprisingly, the second DNA strand transfer during reverse transcription was found to be severely impaired upon stem-loop deletion. Taken together, these results indicate that the stem-loop structure called the dimerization initiation site is a cis element acting on both genomic RNA packaging and synthesis of proviral DNA.},
keywords = {Animals COS Cells DNA, Genetic Virion *Virus Assembly, MARQUET, Non-U.S. Gov't Transcription, Nucleic Acid Support, PAILLART, Post-Translational Proviruses/genetics *RNA, Unité ARN, Viral *Regulatory Sequences, Viral HIV-1/*genetics/physiology Human Mutagenesis Protein Processing, Viral/*biosynthesis Gene Expression Genome},
In retroviruses, the genomic RNA is in the form of a 60S-70S complex composed of two identical genome-length RNA molecules tightly associated through numerous interactions. A major interaction, called the dimer linkage structure, has been found near the RNA 5' end and is probably involved in the control of translation, packaging, and recombination during proviral DNA synthesis. Recently, a small sequence corresponding to a stem-loop structure located in the 5' leader of human immunodeficiency virus type 1 (HIV-1) RNA was found to be required for the initiation of HIV-1 RNA dimerization in vitro and named the dimerization initiation site (E. Skripkin, J.-C. Paillart, R. Marquet, B. Ehresmann, and C. Ehresmann, Proc. Natl. Acad. Sci. USA 91: 4945-4949, 1994). To investigate the possible role of this 5' stem-loop in HIV-1 virion formation and infectivity, four mutant viruses were generated and analyzed in vivo. Results show that deletion of the stem-loop structure reduces infectivity by a factor of 10(3) whereas loop substitutions cause a decrease of 10- to 100-fold. The level of genomic RNA packaging was found to be decreased fivefold in mutants virions containing the stem-loop deletion and only twofold in the loop-substituted virions. Surprisingly, the second DNA strand transfer during reverse transcription was found to be severely impaired upon stem-loop deletion. Taken together, these results indicate that the stem-loop structure called the dimerization initiation site is a cis element acting on both genomic RNA packaging and synthesis of proviral DNA.
Marquet, R; Paillart, J C; Skripkin, E; Ehresmann, C; Ehresmann, B
Localization of the dimerization initiation site of HIV-1 genomic RNA and mechanism of dimerization. Book Chapter
Sarma, R H; Sarma, M H (Ed.): Biological Structure and Dynamics: Proceedings of the Ninth Conversation in the Discipline Biomolecular Stereodynamics, held at the State University of New York at Albany, June 20-24, 1995, 2 , p. 61-72, Adenine Press, 1996.
Liens | BibTeX | Étiquettes: MARQUET, PAILLART, Unité ARN
title = {Localization of the dimerization initiation site of HIV-1 genomic RNA and mechanism of dimerization.},
author = {R Marquet and J C Paillart and E Skripkin and C Ehresmann and B Ehresmann},
editor = {R H Sarma and M H Sarma},
url = {http://books.google.fr/books?hl=fr&id=SOpqAAAAMAAJ&q=Marquet#search_anchor},
booktitle = {Biological Structure and Dynamics: Proceedings of the Ninth Conversation in the Discipline Biomolecular Stereodynamics, held at the State University of New York at Albany, June 20-24, 1995},
publisher = {Adenine Press},
http://books.google.fr/books?hl=fr&id=SOpqAAAAMAAJ&q=Marquet#search_anch[...]
Skripkin, E; Paillart, J C; Marquet, R; Ehresmann, B; Ehresmann, C
Identification of the primary site of the human immunodeficiency virus type 1 RNA dimerization in vitro Article de journal
Résumé | Liens | BibTeX | Étiquettes: Base Sequence Binding Sites Biopolymers HIV-1/*genetics Molecular Sequence Data Nucleic Acid Conformation Nucleotides/chemistry RNA, MARQUET, Non-U.S. Gov't, Nucleic Acid Support, PAILLART, Unité ARN, Viral/*chemistry Repetitive Sequences
title = {Identification of the primary site of the human immunodeficiency virus type 1 RNA dimerization in vitro},
author = {E Skripkin and J C Paillart and R Marquet and B Ehresmann and C Ehresmann},
abstract = {The diploid genome of all retroviruses is made of two homologous copies of RNA intimately associated near their 5' end, in a region called the dimer linkage structure. Dimerization of genomic RNA is thought to be important for crucial functions of the retroviral life cycle (reverse transcription, translation, encapsidation). Previous in vitro studies mapped the dimer linkage structure of human immunodeficiency virus type 1 (HIV-1) in a region downstream of the splice donor site, containing conserved purine tracts that were postulated to mediate dimerization, through purine quartets. However, we recently showed that dimerization of HIV-1 RNA also involves sequences upstream of the splice donor site. Here, we used chemical modification interference to identify nucleotides that are required in unmodified form for dimerization of a RNA fragment containing nucleotides 1-707 of HIV-1 RNA. These nucleotides map exclusively in a restricted area upstream of the splice donor site and downstream of the primer binding site. They are centered around a palindromic sequence (GUGCAC279) located in a hairpin loop. Our results support a model in which dimer formation is initiated by the annealing of the palindromic sequences, possibly by a loop-loop interaction between the two monomers. Further experiments show that the deletion of the stem-loop or base substitutions in the loop abolish dimerization, despite the presence of the previously postulated dimer linkage structure. On the other hand, deletions of the purine tracts downstream of the splice donor site do not prevent dimerization. Therefore, we conclude that the palindromic region represents the dimerization initiation site of genomic RNA.},
keywords = {Base Sequence Binding Sites Biopolymers HIV-1/*genetics Molecular Sequence Data Nucleic Acid Conformation Nucleotides/chemistry RNA, MARQUET, Non-U.S. Gov't, Nucleic Acid Support, PAILLART, Unité ARN, Viral/*chemistry Repetitive Sequences},
The diploid genome of all retroviruses is made of two homologous copies of RNA intimately associated near their 5' end, in a region called the dimer linkage structure. Dimerization of genomic RNA is thought to be important for crucial functions of the retroviral life cycle (reverse transcription, translation, encapsidation). Previous in vitro studies mapped the dimer linkage structure of human immunodeficiency virus type 1 (HIV-1) in a region downstream of the splice donor site, containing conserved purine tracts that were postulated to mediate dimerization, through purine quartets. However, we recently showed that dimerization of HIV-1 RNA also involves sequences upstream of the splice donor site. Here, we used chemical modification interference to identify nucleotides that are required in unmodified form for dimerization of a RNA fragment containing nucleotides 1-707 of HIV-1 RNA. These nucleotides map exclusively in a restricted area upstream of the splice donor site and downstream of the primer binding site. They are centered around a palindromic sequence (GUGCAC279) located in a hairpin loop. Our results support a model in which dimer formation is initiated by the annealing of the palindromic sequences, possibly by a loop-loop interaction between the two monomers. Further experiments show that the deletion of the stem-loop or base substitutions in the loop abolish dimerization, despite the presence of the previously postulated dimer linkage structure. On the other hand, deletions of the purine tracts downstream of the splice donor site do not prevent dimerization. Therefore, we conclude that the palindromic region represents the dimerization initiation site of genomic RNA.


text:
Originally published: 2001-06-01 07:53:48
Last updated: 2001-06-01 07:53:48
Author: alan runyan

has a base Factory class that is meant to be subclassed and then define a default implementation to return, as well as module's to search for the class implementation.

redpajama_set_name: RedPajamaGithub


Each scene in the drama of salvation had its own timing enabling God to both be just and the justifier in our behalf. In this concise study, various aspects of Christ's ministry that are key to providing salvation for mankind are explored. This will refresh your faith and provide a deeper understanding of His great atoning sacrifice.


text:
April 27, 2014 / 9:59 AM / 6 years ago
Siemens and France weigh in as GE eyes Alstom deal
Benjamin Mallet, Maria Sheahan
PARIS/FRANKFURT (Reuters) - Siemens and the French government intervened in General Electric's plan to buy the power arm of Alstom on Sunday with an alternative European "champions" tie-up proposal and a pledge to act in France's national interest.
View of a Haliade 150 offshore wind turbine at Alstom's offshore wind site in Le Carnet, on the Loire Estuary, near Saint Nazaire, western France, April 27, 2014. REUTERS/Stephane Mahe
Though French trains-to-turbines maker Alstom is privately owned, firebrand Economy Minister Arnaud Montebourg issued a stark reminder of the influence the government holds over a company that relies heavily on orders from state rail operator SNCF and partly state-owned utility EDF.
"GE and Alstom have their calendar, which is that of shareholders, but the French government has its own, which is that of economic sovereignty," Montebourg said in a statement, providing the first official confirmation of GE's offer.
GE boss Jeff Immelt was in Paris on Sunday to thrash out a $13 billion (7 billion pounds) deal for struggling Alstom's power turbines and grid equipment. Montebourg had planned to meet him but the encounter was postponed until later in the week after the minister advertised an alternative proposal by German rival Siemens.
The Siemens proposal would create "two European and global champions in the energy and transport domains - one around Siemens, the other around Alstom", Montebourg said.
French President Francois Hollande gathered his top ministers on Sunday evening to discuss Alstom's case. The company said separately it would make an announcement no later than Wednesday morning and that it had asked for its shares to remain suspended from trading until then.
"Alstom continues and deepens its strategic reflection," it said in a three-line statement.
Montebourg said the government would not accept any hastily made decision, that it would seek to preserve France's jobs and industrial base and would in particular be "extremely vigilant" in ensuring the nation's nuclear industry remains independent.
The warning compels Alstom and GE to tread carefully. However, sources familiar with the talks said these were very advanced.
"Alstom has received a firm offer from GE and an expression of interest from Siemens. It's not at all the same kind of commitment," one of the sources said.
TRAINS PLUS CASH
Earlier in the day, Siemens entered the fray with an announcement that it had written a letter to "signal its willingness to discuss future strategic opportunities" with the French group. Siemens gave no further details.
Hollande to meet GE chief on Monday to discuss Alstom
French president, GE boss to meet over Alstom future
A report on newspaper Le Figaro's website on Sunday said Siemens was offering Alstom half of its train-making business plus cash in exchange for its French rival's power turbines division.
Le Figaro said Siemens was proposing that Alstom take on the Siemens high-speed trains and locomotives arm, but not its metropolitan trains division. A report in Germany's Handelsblatt outlined a similar scenario and put the value of the proposed deal at 10-11 billion euros ($14-15 billion).
Alstom CEO Patrick Kron has said in the past he is against creating a Franco-German train manufacturer.
As well as the power turbines arm GE wants to buy, Alstom makes TGV high-speed trains and is one of France's top private-sector employers.
Though Alstom is struggling with heavy debt and weak demand, political sensitivities run deep in France.
Marine Le Pen, leader of the far-right National Front (FN) party that won widespread support in last month's local elections, said the government had "abandoned Alstom to be dismantled for American or German profit".
A Siemens tie-up may be no more palatable to some than a deal with GE. A decade ago, Alstom was rescued by a state-backed restructuring when Kron and France's then-president, Nicolas Sarkozy, both balked at the prospect of a Siemens acquisition.
A deal to sell Alstom's power assets, which account for about 70 percent of total group revenue, would effectively break up the engineering group and leave Alstom as a pure transport business, building its TGV trains, other rolling stock and rail signalling equipment.
Alstom's power assets include turbines for coal, gas and nuclear plants, wind farms and systems for power transmission and distribution. They generated around 15 billion euros or 70 percent of Alstom's global revenue in the last fiscal year and are among the group's most profitable businesses.
Alstom is the world's top supplier of turbine generator sets for nuclear plants. The company estimates that its equipment, which is used to produce electricity but is not located inside nuclear reactors, is used in 40 percent of nuclear plants.
Alstom employs 18,000 people in France, about 20 percent of its total workforce, against GE's 10,000 French workers and Siemens' 7,000. A source familiar with the matter said there was lots of overlap between Siemens and Alstom's operations in France, more so than with GE.
Alstom and Siemens are direct rivals in steam turbines, offshore wind power, hydro power and grid. For its part, GE is absent from offshore wind and hydro power and could use the boost in steam turbines and grid.
Sources have said a deal with GE is backed by Alstom's main shareholder, French conglomerate Bouygues, which holds a 29 percent stake.
Before the news of an approach from GE, Alstom shares had slumped 20 percent in 12 months on concerns over its cash flow, prompting Bouygues to take a $1.9 billion writedown on its stake in February. Trading of Alstom shares was suspended on Friday at the request of market regulator AMF.
Additional reporting by Andrew Callus, Matthieu Protard and Natalie Huet in Paris, Anjuli Davies in London and Soyoung Kim in New York; Editing by David Goodman and Gareth Jones

redpajama_set_name: RedPajamaCommonCrawl


Min Qty "PharmaHemp CBD Oil Drops 5% CBD in 10 ml bottle" is 1.
Hemp grown in the lush organic farmlands of Slovenia and then harvested by hand, this blend 5% CBD oil from PharmaHemp sets the trend for quality CBD oils. Extracted using steam distillation to retain all its nutrients, is 100% organic and is GMO FREE!, it' easy to see why this oil is so popular and sought after. Unlike other brands that have jumped on the "CBD bandwagon", PharmaHemp have always used ethical hemp growing methods, lets not forget the company have been around since 1965 so have had plenty of time to adapt their products.
PharmaHemp CBD oil drops are available in 5 different grades of CBD oil. To preserve all the beneficial compounds in our extracts, PharmaHemp uses highly efficient supercritical CO2 extraction, which leads to a high purity of quality CBD. Pure organic with NO NASTIES and made with lots of love, Pharma Hemp 5% CBD oil, also containing high levels of fatty acids and vitamins. The 5% CBD content across all our brands is one of the most popular sold by The Hemp Doctor and being totally natural with the powerful rejuvenating properties of CBD, this oil is a "Must Try".
Pharma-Hemp's 5% CBD oil is a great way to introduce a totally natural supplement to enhance your health and well-being and it's perfect for everyday use.


text:
import * as ReduxUtils from '../core-utils/redux-utils';
import { Actions } from '../actions';
import GD from '../constants';
import { PecaMimeType } from '../core-utils/file-utils';
import * as stringUtils from '../core-utils/string-utils';

const { ActionsSuffix } = ReduxUtils;

export function autosReducer(state, action) {
    if (state === undefined) {
        state = stateDefault();
    }
    switch(action.type) {
    case Actions.AUTOS_CLEAR_STATE:
        return stateDefault();

    case Actions.AUTOS_SET_PECAS:
        return setPecas(action, state);

    case Actions.AUTOS_ADD_PECA:
        return addPeca(action, state);

    case Actions.AUTOS_SELECT_PECA:
        return selectPeca(action, state);

    case Actions.AUTOS_SELECT_PECA_PAGINA:
        return selectPecaPagina(action, state);

    case Actions.AUTOS_SET_PECA_PDF_URL:
        return setPecaPdfUrl(action, state);

    case Actions.AUTOS_FETCH_PECA_CONTEUDO_HTML + ActionsSuffix.SUCCESS:
        return fetchPecaConteudoHtmlSuccess(action, state);

    case Actions.AUTOS_DELETE_MARCACAO:
        return deleteMarcacao(action, state);

    case Actions.AUTOS_CREATE_MARCACAO + ActionsSuffix.SUCCESS:
        return createMarcacaoSuccess(action, state);

    case Actions.AUTOS_SELECT_MARCACAO:
        return selectMarcacao(action, state);

    case Actions.AUTOS_FETCH_MOVIMENTACOES + ActionsSuffix.SUCCESS:
        return fetchMovimentacoes(action, state);

    case Actions.AUTOS_REMOVE_PECA:
        return removePeca(action, state);

    case Actions.AUTOS_SELECT_FIRST_PECA:
        return selectFirstPeca(action, state);

    case Actions.AUTOS_SET_MARCACAO_SELECIONADA:
        return setMarcacaoSelecionada(action, state);

    case Actions.AUTOS_SCROLL_TO_MARCACAO:
        return scrollToMarcacao(action, state);

    case Actions.AUTOS_ACCEPT_MARCACAO + ActionsSuffix.SUCCESS:
        return acceptMarcacaoSuccess(action, state);

    case Actions.AUTOS_REJECT_MARCACAO + ActionsSuffix.SUCCESS:
        return rejectMarcacaoSuccess(action, state);

    case Actions.AUTOS_START_LINK:
        return startLink(action, state);

    case Actions.AUTOS_CANCEL_LINK:
        return cancelLink(action, state);

    case Actions.AUTOS_CONCLUIR_LINK + ActionsSuffix.SUCCESS:
        return concluirLinkSuccess(action, state);

    case Actions.AUTOS_ADD_PAGINA_TO_LINK:
        return addPaginaToLink(action, state);

    case Actions.AUTOS_GO_LINK + ActionsSuffix.SUCCESS:
        return goLinkSuccess(action, state);

    case Actions.AUTOS_CLOSE_LINK_BAR:
        return closeLinkBar(action, state);

    case Actions.AUTOS_BACK_TO_LINK_SUCCESS:
        return backToLinkSuccess(action, state);

    case Actions.AUTOS_CLEAR_ACTIVE_LINK_TO_SCROLL:
        return clearActiveLinkToScroll(action, state);

    case Actions.AUTOS_ZOOM_IN:
        return zoomIn(state);

    case Actions.AUTOS_ZOOM_OUT:
        return zoomOut(state);

    case Actions.AUTOS_ACTIVE_PAGE:
        return activePage(action, state);

    case Actions.AUTOS_START_PECA_EDIT:
        return startPecaEdit(action, state);

    case Actions.AUTOS_SAVE_PECA + ActionsSuffix.SUCCESS:
        return savePecaSuccess(action, state);

    case Actions.AUTOS_DELETE_LINK_SUCCESS:
        return deleteLinkSuccess(action, state);

    case Actions.AUTOS_CREATE_PECA_AVULSA + ActionsSuffix.SUCCESS:
        return createPecaAvulsaSuccess(action, state);

    case Actions.AUTOS_UPDATE_PECA_AVULSA:
        return updatePecaAvulsa(action, state);

    default:
        return state;
    }
}

function stateDefault() {
    return {
        isFetching: false,
        pecas: [],
        pecasById: {},
        pecaSelecionada: {},
        navContracted: false,
        error: undefined,
        movimentacoes: [],
        pecaConteudo: {
            mimeType: null,
            conteudo: null
        },
        marcacaoSelecionada: null,
        marcacaoScroll: null,
        linkEmConstrucao: null,
        numeroPaginaSelecionada: 1,
        zoom: 1,
        activePage: 1,
        activeLink: null,
        activeLinkPecaId: null,
        pecaEditingId: null,
        pecaAvulsa: null
    };
}

function setPecas(action, state) {
    const pecas = [...action.pecas];
    const pecasById = getElementosById(pecas);
    return {
        ...state,
        pecas: pecas,
        pecasById: pecasById
    };
}

function addPeca(action, state) {
    const pecas = [...state.pecas];
    pecas.push(action.peca);
    const pecasById = {...state.pecasById};
    pecasById[action.peca.id] = action.peca;
    return {
        ...state,
        pecas: pecas,
        pecasById: pecasById
    };
}

function selectPeca(action, state) {
    const peca = state.pecasById[action.pecaId];
    return {
        ...state,
        pecaSelecionada: {...peca},
        numeroPaginaSelecionada: 1,
    };
}

function selectPecaPagina(action, state) {
    const peca = state.pecasById[action.pecaId];
    return {
        ...state,
        pecaSelecionada: {...peca},
        numeroPaginaSelecionada: action.numeroPagina,
        activePage: action.numeroPagina
    };
}

function setPecaPdfUrl(action, state) {
    let peca = state.pecasById[action.pecaId];
    peca = {...peca};
    if (peca.mimeType === PecaMimeType.PDF) {
        let url = GD.BACKEND + '/service/processo/:processoId/peca/:pecaId/pdf';
        url = url.replace(':processoId', action.processoId).replace(':pecaId', peca.id);
        peca.url = url;
    }
    return {
        ...state,
        pecaSelecionada: peca
    };
}

function fetchPecaConteudoHtmlSuccess(action, state) {
    const pecasById = {...state.pecasById};
    let pecaSelecionada = pecasById[action.pecaId];
    const index = state.pecas.indexOf(pecaSelecionada);
    pecaSelecionada = {...pecaSelecionada};
    pecaSelecionada.conteudo = action.data;
    const pecas = [...state.pecas];
    pecas[index] = pecaSelecionada;
    pecasById[action.pecaId] = pecaSelecionada;
    return {
        ...state,
        pecas,
        pecasById,
        pecaSelecionada
    };
}

function deleteMarcacao(action, state) {
    let newState = {...state};
    let pecas = ReduxUtils.cloneProperty(newState, 'pecas');
    let peca = ReduxUtils.cloneItemArrayById(pecas, action.pecaId);
    let marcacoes = ReduxUtils.cloneProperty(peca, 'marcacoes');
    ReduxUtils.deleteItemArrayById(marcacoes, action.marcacaoId);
    let pecaSelecionada = ReduxUtils.cloneProperty(newState, 'pecaSelecionada');
    if (pecaSelecionada.id === peca.id) {
        pecaSelecionada.marcacoes = marcacoes;
    }
    let pecasById = ReduxUtils.cloneProperty(newState, 'pecasById');
    pecasById[action.pecaId] = peca;
    return newState;
}

function createMarcacaoSuccess(action, state) {
    let newState = {...state};
    const marcacao = action.data.marcacao;

    let pecaSelecionada = ReduxUtils.cloneProperty(newState, 'pecaSelecionada');
    let marcacoes = ReduxUtils.cloneProperty(pecaSelecionada, 'marcacoes');
    marcacoes.push(marcacao);
    pecaSelecionada.marcacoes = marcacoes;
    newState.marcacaoSelecionada = marcacao;
    newState.pecaSelecionada = pecaSelecionada;

    let pecas = ReduxUtils.cloneProperty(newState, 'pecas');
    let peca = ReduxUtils.cloneItemArrayById(pecas, action.data.peca.id);
    const index = pecas.indexOf(peca);
    pecas[index] = pecaSelecionada;

    newState.pecas = pecas;

    let pecasById = ReduxUtils.cloneProperty(newState, 'pecasById');
    pecasById[action.data.peca.id] = pecaSelecionada;

    newState.pecasById = pecasById;

    return newState;
}

function selectMarcacao(action, state) {
    const marcacoes = state.pecaSelecionada.marcacoes;
    const marcacaoSelecionada = ReduxUtils.findItemById(marcacoes, action.marcacaoId);
    return {
        ...state,
        marcacaoSelecionada
    };
}

function fetchMovimentacoes(action, state) {
    return {
        ...state,
        movimentacoes: action.data
    };
}

function removePeca(action, state) {
    const peca = ReduxUtils.findItemById(state.pecas, action.pecaId);
    const indexPeca = state.pecas.indexOf(peca);
    const pecas = [...state.pecas];
    pecas.splice(indexPeca, 1);
    return {
        ...state,
        pecas
    };
}

function selectFirstPeca(action, state) {
    const peca = {...state.pecas[0]};
    return {
        ...state,
        pecaSelecionada: peca
    };
}

function setMarcacaoSelecionada(action, state) {
    const pecaSelecionada = state.pecaSelecionada;
    const marcacaoSelecionada = ReduxUtils.findItemById(pecaSelecionada.marcacoes, action.marcacaoId);
    return {
        ...state,
        marcacaoSelecionada
    };
}

function scrollToMarcacao(action, state) {
    const pecaSelecionada = state.pecaSelecionada;
    const marcacaoScroll = ReduxUtils.findItemById(pecaSelecionada.marcacoes, action.marcacaoId);
    return {
        ...state,
        marcacaoScroll
    };
}

function acceptMarcacaoSuccess(action, state) {
    let { marcacaoId, pecaId } = action;

    let newState = {...state};
    let pecas = ReduxUtils.cloneProperty(newState, 'pecas');
    let peca = ReduxUtils.cloneItemArrayById(pecas, pecaId);
    let marcacoes = ReduxUtils.cloneProperty(peca, 'marcacoes');
    let marcacao = ReduxUtils.cloneItemArrayById(marcacoes, marcacaoId);
    marcacao.sugerida = false;

    peca.marcacoes = marcacoes;
    newState.pecaSelecionada = peca;
    let pecasById = ReduxUtils.cloneProperty(newState, 'pecasById');
    pecasById[action.pecaId] = peca;
    return newState;
}

function rejectMarcacaoSuccess(action, state) {
    let { marcacaoId, pecaId } = action;

    let newState = {...state};
    let pecas = ReduxUtils.cloneProperty(newState, 'pecas');
    let peca = ReduxUtils.cloneItemArrayById(pecas, pecaId);
    let marcacoes = ReduxUtils.cloneProperty(peca, 'marcacoes');
    ReduxUtils.deleteItemArrayById(marcacoes, marcacaoId);

    peca.marcacoes = marcacoes;
    newState.pecaSelecionada = peca;
    let pecasById = ReduxUtils.cloneProperty(newState, 'pecasById');
    pecasById[action.pecaId] = peca;

    newState.marcacaoSelecionada = null;

    return newState;
}

function startLink(action, state) {
    const linkEmConstrucao = {
        pecaId: state.pecaSelecionada.id,
        selecao: action.selecao,
        totalAlvos: 0
    };
    return {
        ...state,
        linkEmConstrucao
    };
}

function cancelLink(action, state) {
    removePaginasEmConstrucao(state);
    const pecas = [...state.pecas];
    return {
        ...state,
        linkEmConstrucao: null,
        pecas
    };
}

function concluirLinkSuccess(action, state) {
    const linkEmConstrucao = state.linkEmConstrucao;
    const pecaId = linkEmConstrucao.pecaId;
    const peca = state.pecasById[pecaId];
    const link = action.data;
    peca.links.push(link);
    confirmPaginasEmConstrucao(state);
    const pecas = [...state.pecas];
    return {
        ...state,
        linkEmConstrucao: null,
        activeLinkToScroll: link,
        pecas
    };
}

function addPaginaToLink(action, state) {
    const linkEmConstrucao = {...state.linkEmConstrucao};
    const pecaSelecionada = {...state.pecaSelecionada};
    if (!pecaSelecionada.paginasAlvo) {
        pecaSelecionada.paginasAlvo = {};
    } else {
        pecaSelecionada.paginasAlvo = {...pecaSelecionada.paginasAlvo};
    }

    if (pecaSelecionada.paginasAlvo[action.numero]) {
        pecaSelecionada.paginasAlvo[action.numero].emConstrucao = true;
    } else {
        const descricao = stringUtils.upperFirst(linkEmConstrucao.selecao.conteudo);
        pecaSelecionada.paginasAlvo[action.numero] = {
            descricao,
            emConstrucao: true
        };
    }

    const index = getIndexPecaSelecionada(state);
    const pecas = [...state.pecas];
    pecas[index] = pecaSelecionada;
    const pecasById = getElementosById(pecas);

    linkEmConstrucao.totalAlvos++;

    return {
        ...state,
        linkEmConstrucao,
        pecaSelecionada,
        pecas,
        pecasById
    };
}

function goLinkSuccess(action, state) {
    const { link } = action;
    const firstAlvo = link.alvos[0];
    addPaginaInicialToAlvos(state, link.alvos);
    return {
        ...state,
        activeLink: link,
        activeLinkPecaId: action.pecaId,
        numeroPaginaSelecionada: firstAlvo.pagina,
        activePage: firstAlvo.pagina,
    };
}

function closeLinkBar(action, state) {
    return {
        ...state,
        activeLink: null,
        activeLinkPecaId: null
    };
}

function backToLinkSuccess(action, state) {
    const pecaId = state.activeLinkPecaId;
    const pecasById = {...state.pecasById};
    const pecaSelecionada = {...pecasById[pecaId]};

    return {
        ...state,
        activeLink: null,
        activeLinkToScroll: state.activeLink,
        activeLinkPecaId: null,
        pecaSelecionada
    };
}

function clearActiveLinkToScroll(action, state) {
    return {
        ...state,
        activeLinkToScroll: null
    };
}

function zoomIn(state) {
    let zoom = state.zoom += 0.1;
    zoom = zoom < 5 ? zoom : 5;
    return {
        ...state,
        zoom
    };
}

function zoomOut(state) {
    let zoom = state.zoom -= 0.1;
    zoom = zoom > 0.2 ? zoom : 0.2;
    return {
        ...state,
        zoom
    };
}

function activePage(action, state) {
    return {
        ...state,
        activePage: action.page
    };
}

function startPecaEdit(action, state) {
    return {
        ...state,
        pecaEditingId: action.pecaId,
    };
}

function savePecaSuccess(action, state) {
    const pecaEdited = findPecaById(state, state.pecaEditingId);
    pecaEdited.descricao = action.descricao;
    const pecas = [...state.pecas];
    const index = pecas.indexOf(pecaEdited);
    pecas[index] = {...pecaEdited};
    const pecasById = getElementosById(pecas);
    return {
        ...state,
        pecaEditingId: null,
        pecas,
        pecasById
    };
}

function deleteLinkSuccess(action, state) {
    const pecasAlvosDeletados = deletePaginaAlvo(action, state);
    const pecasLinksDeletados = deletePecaLink(pecasAlvosDeletados, state);

    return {
        ...state,
        'pecas': pecasLinksDeletados,
        'pecasById': getElementosById(pecasLinksDeletados),
        activeLink: null
    };
}

function createPecaAvulsaSuccess(action, state) {
    const pecaAvulsa = action.pecaAvulsa;
    const peca = {
        id: action.pecaAvulsa.id,
        descricao: 'Peca em Elaboração',
        polo: 'JUIZ',
        principal: true,
        data: Date.now(),
        marcacoes: []
    };
    const pecas = [...state.pecas];
    pecas.push(peca);
    return {
        ...state,
        pecaAvulsa,
        pecas
    };
}

function updatePecaAvulsa(action, state) {
    const pecaAvulsa = action.pecaAvulsa;
    return {
        ...state,
        pecaAvulsa,
    };
}

function deletePaginaAlvo(action, state) {
    const alvosIdToDelete = action.data;
    const pecas = [];

    for (const peca of state.pecas) {
        let pecaAlterada = false;

        for (const pagina in peca.paginasAlvo) {
            const alvo = peca.paginasAlvo[pagina];

            if (alvosIdToDelete.includes(alvo.id)) {
                delete peca.paginasAlvo[pagina];
                pecaAlterada = true;
            }
        }

        if (pecaAlterada) {
            let pecaClone = {...peca};
            pecas.push(pecaClone);
        } else {
            pecas.push(peca);
        }
    }
    return pecas;
}

function deletePecaLink(pecas, state) {
    const peca = pecas.find(p => p.id === state.activeLinkPecaId);

    for (let i = 0; i < peca.links.length; i++) {
        const link = peca.links[i];

        if (link.id === state.activeLink.id) {
            peca.links.splice(i, 1);
        }
    }

    return pecas;
}

// uteis ====================

function getElementosById(elementos) {
    let elementosById = {};
    for (let i = 0; i < elementos.length; i++) {
        let elemento = elementos[i];
        elementosById[elemento.id] = elemento;
    }
    return elementosById;
}

function getIndexPecaSelecionada(state) {
    const pecaSelecionadaId = state.pecaSelecionada.id;
    for (let i = 0; i < state.pecas.length; i++) {
        if (state.pecas[i].id === pecaSelecionadaId) {
            return i;
        }
    }
    throw {
        type: 'ERROR',
        title: 'Não existe peça selecionada'
    };
}

function removePaginasEmConstrucao(state) {
    for (let i = 0; i < state.pecas.length; i++) {
        const peca = state.pecas[i];
        if (!peca.paginasAlvo) {
            continue;
        }
        for (const numero in peca.paginasAlvo) {
            const pagina = peca.paginasAlvo[numero];
            if (pagina.emConstrucao) {
                if (pagina.id) {
                    delete pagina.emConstrucao;
                } else {
                    delete peca.paginasAlvo[numero];
                }
            }
        }
    }
}

function confirmPaginasEmConstrucao(state) {
    for (let i = 0; i < state.pecas.length; i++) {
        const peca = state.pecas[i];
        if (!peca.paginasAlvo) {
            continue;
        }
        for (const numero in peca.paginasAlvo) {
            const pagina = peca.paginasAlvo[numero];
            if (pagina.emConstrucao) {
                pagina.emConstrucao = false;
                delete pagina.emConstrucao;
            }
        }
    }
}

function addPaginaInicialToAlvos(state, alvos) {
    const pecasById = state.pecasById;
    alvos.forEach(item => {
        item.paginaInicial = pecasById[item.pecaId].paginaInicial;
    });
}

function findPecaById(state, pecaId) {
    for (let i = 0; i < state.pecas.length; i++) {
        const peca = state.pecas[i];
        if (peca.id === pecaId) {
            return peca;
        }
    }
    return null;
}