You will be presented with the abstract, introduction, and discussion section from a biomedical research article. Your task is to create a summary that a layperson can understand, capturing the essence of the research article. Keep essential scientific terms, but ensure that the language remains clear and the concepts are explained in an uncomplicated manner.

Title: Infection with flaviviruses requires BCLXL for cell survival}
Article:
Abstract:
BCL2 family proteins including pro-survival proteins, BH3-only proteins and BAX/BAK proteins control mitochondria-mediated apoptosis to maintain cell homeostasis via the removal of damaged cells and pathogen-infected cells. In this study, we examined the roles of BCL2 proteins in the induction of apoptosis in cells upon infection with flaviviruses, such as Japanese encephalitis virus, Dengue virus and Zika virus. We showed that survival of the infected cells depends on BCLXL, a pro-survival BCL2 protein due to suppression of the expression of another pro-survival protein, MCL1. Treatment with BCLXL inhibitors, as well as deficient BCLXL gene expression, induced BAX/BAK-dependent apoptosis upon infection with flaviviruses. Flavivirus infection attenuates cellular protein synthesis, which confers reduction of short-half-life proteins like MCL1. Inhibition of BCLXL increased phagocytosis of virus-infected cells by macrophages, thereby suppressing viral dissemination and chemokine production. Furthermore, we examined the roles of BCLXL in the death of JEV-infected cells during in vivo infection. Haploinsufficiency of the BCLXL gene, as well as administration of BH3 mimetic compounds, increased survival rate after challenge of JEV infection and suppressed inflammation. These results suggest that BCLXL plays a crucial role in the survival of cells infected with flaviviruses, and that BCLXL may provide a novel antiviral target to suppress propagation of the family of Flaviviridae viruses.
Introduction:
The genus Flavivirus belongs to the family Flaviviridae and includes the Japanese encephalitis virus( JEV), Dengue virus( DENV), West Nile virus( WNV) and Zika virus( ZIKV), all of which are mosquito-borne human pathogens[1, 2]. The flaviviruses are internalized into dendritic cells, such as Langerhans cells[3], and keratinocytes in skin[4] through mosquito bites. Dendritic cells that are thus infected with flaviviruses are activated and migrate into the lymph nodes, facilitating viral spread into peripheral tissues[5] and the induction of visceral and/or central nervous system diseases. DENV infection may generate dengue haemorrhagic fever and dengue shock syndrome[6], while infection with either WNV or JEV can cause severe encephalitis[7, 8]. Recently, an outbreak of ZIKV infection in Brazil was linked to birth defects, including microcephaly, and to Guillain-Barre syndrome in adults[9, 10]. Therefore, the development of effective vaccines and antiviral drugs against the family of flaviviruses is an urgent need. Infection with flaviviruses induces apoptosis in cells by the production of reactive oxidative species, unfolded protein responses, and viral proteins[11], in addition to other types of cell death, such as necrosis and pyroptosis. Receptor-interacting serine/threonine-protein kinase-3( RIPK3), a central player of necroptosis, has been shown to play an additional role during virus infection[12]. Infection with either DENV or hepatitis C virus( HCV) induces interleukin-1β production and pyroptosis[13]. The biological significance of pyroptosis in the pathogenicity of flavivirus infection remains largely unknown[14]. Apoptosis is a type of programmed cell death that removes excess cells during development, as well as during routine homeostasis[15–17]. B-cell lymphoma-2( BCL2) family proteins play central roles in mitochondria-mediated apoptosis[18]. BCL2 family proteins have been divided into three groups: BH3-only proteins, BAX/BAK proteins and pro-survival proteins. Once an apoptotic stimulus such as DNA damage occurs, BH3-only proteins, such as BIM, BID, PUMA, BAD and NOXA, are activated and directly or indirectly activate BAX and BAK. Activated BAX/BAK forms homo-oligomers that permeabilize mitochondria and allow the release of pro-apoptotic factors such as cytochrome c, which promotes activation of caspases[19]. Pro-survival proteins, such as BCL2, BCLW, BCLXL, MCL1 and A1, sequester BH3-only proteins to prevent self-activation of BAX/BAK by interaction with the BH3 domains of BAX/BAK[20]. In a notable extension to cancer therapy, small molecule inhibitors have been developed, including ABT-737, ABT-263( navitoclax, an orally available clinical derivative of ABT-737), ABT-199( venetoclax) and A-1331852. ABT-199 and A-1331852 target BCL2 and BCLXL, respectively, while ABT-737 and ABT-263 target BCL2, BCLXL and BCLW. ABT-199 was approved by the United States Food and Drug Administration for the treatment of 17-p deleted chronic lymphocytic leukaemia( CLL)[21], supporting the therapeutic use of induction of cell death by mitochondria-mediated apoptosis to eliminate unnecessary cells. In this study, we showed that cells infected with flaviviruses including JEV, DENV and ZIKV specifically induced apoptosis by inhibition of BCLXL through the suppression of MCL1 expression. Furthermore, we showed that host protein synthesis is impaired in cells upon infection with flaviviruses, leading to the decay of labile proteins such as MCL1. Treatment with inhibitors for BCLXL, as well as deficiency of BCLXL gene, induced BAX/BAK-dependent apoptosis in the infected cells. Finally, engulfment of virus-infected cells by macrophages was significantly enhanced by the inhibition of BCLXL without an increase of the production of pro-inflammatory cytokines, which led to the suppression of viral dissemination.
Discussion:
In this study, we showed that flavivirus infection suppresses expression of labile proteins such as MCL1 without observable cytotoxicity. Clear cell death, through activation of caspase 3/7, occurred only when BCLXL was inhibited. Moreover, we showed that treatment with BCLXL inhibitor( A-1331852) or deficiency of the BCLXL gene induces cell death upon infection with flaviviruses in several cancer cell lines and primary MEFs. Previous report showed that BCLXL and MCL1 predominantly inhibit activation of BAK[50], suggesting that a mechanism to inhibit BCLXL may induce BAK-dependent apoptosis in flavivirus-infected cells, therefore inhibition of BCLX alone could enhanced cell death in virus-infected cells. In addition, viral infection decreased MCL1 expression in a variety of human and murine cell lines. Although the magnitude of suppression of MCL1 expression varied depending on cell line, our data suggest that suppression of MCL1 expression upon propagation of flaviviruses is a common feature. Legionella infection also induces suppression of MCL1 expression[51]. Although the molecular mechanisms of MCL1 protein expression in cells infected with Legionella remain unclear, the suppression of MCL1 through shut off of host translation might be part of the cellular innate response against infection with intracellular pathogens. MCL1 expression is strictly controlled by transcriptional and post-transcriptional mechanisms[52]. In this study, we showed that flavivirus infection suppresses protein synthesis, thereby reducing the quantity of proteins with short half-lives, such as MCL1. In addition, recent reports have shown that infection with flaviviruses represses translation of host proteins, but does not repress viral proteins, which are independent of PKR and eIF2α[53]. Further, infection with mammalian orthoreovirus has been shown to inhibit host translation by compartmentalization of translation machinery within the viral replication complex[54]. Protein translation is regulated by ribosomal proteins[55]. DENV NS1 protein specifically binds to the ribosomal protein RPL18 to sequester in perinuclear region and to regulate viral translation and replication[56]. RPLP1 and RPLP2 identified by genome-wide RNAi screening were host factors to contribute DENV and yellow fever virus( YFV) infection[57]. We previously reported that heterogeneous nuclear ribonucleoprotein A2/B1( HNRNPA2B1) interact with JEV core protein and facilitate JEV replication through the translocation of core protein from nucleus to cytoplasm[58]. In addition, infection with HCV has been shown to induce accumulation of adenosine 5′-triphosphate in the viral replication complex[59]. Because expression of JEV proteins exhibited no effect on host translation( S3L Fig), viral propagation might induce shut-off of host translation by unknown mechanism rather than PKR and eIF2α. On the other hand, viruses counteract host responses through the rearrangement of membrane structures to restrict the components of translation machinery into the viral replication complex. Our data indicate that inhibition of BCLXL in viral-infected cells activates phagocytosis by macrophages, leading to the suppression of infectious particle production and suggest that inhibition of BCLXL function as an anti-viral effect. Although the survival of mice that were intracerebrally inoculated with JEV was not significantly different between wild-type and Bclx+/- mice, Bclx+/- mice showed higher survival rate against subcutaneous challenge by JEV, compared with their wild-type counterparts. Our data suggest that phagocytosis plays an important role in removing apoptotic. JEV-infected cells after BCLX inhibition. Microglial cells, a resident subset of macrophages in the brain, and hematogenous macrophages play a role in phagocytosis in the brain and peripheral tissues, respectively. These two-types of macrophage have phagocytotic activity but have some different functions[60]. The functional difference of the microglia and hematogenous macrophages might be involved in the phagocytosis-mediated removals of virus-infected cells by BCLX inhibition in intracerebral or subcutaneous infection. Collectively, these data suggest that apoptosis induced by inhibition of BCLXL attenuates viral virulence in peripheral tissues. The production of cytokines and chemokines during viral infection is strongly associated with viral pathogenicity[61], especially, deficiency of several chemokine receptors such as CCR5 and CXCR3 promotes severity of pathogenicity against WNV infection[62]. We showed that the inhibition of BCLXL specifically induces apoptosis and impaired production of CCL5 and CXCL10, which are ligands for CCR5 and CXCR3, respectively, during in vitro and in vivo viral infections. Impairment of these chemokines may contribute attenuation of pathogenicity by inhibition of BCLXL. In addition, administration of the BH3 mimetic, ABT-263, into wild-type mice increased survival rate after subcutaneous JEV challenge. We also showed that production of viral RNA and chemokines was impaired in Bclx+/- mice( Fig 6B and 6C). In addition, treatment of wild type and Bclx+/- mice with polyIC exhibited no effect on chemokine induction( Fig 6D). These data suggest that enhanced apoptosis in Bclx+/- and ABT-263-treated mice upon infection with JEV promotes engulfment by phagocytes, which leads to reduces viral production and the number of leukocytes migrating to the infection site due to lower production of chemokines. Treatment with inhibitors of BCLX such as ABT-263 was demonstrated to cause thrombocytopenia due to their neutralization of BCLXL in circulating platelets without bone marrow toxicity[30, 63, 64]. It was also demonstrated that thrombocytopenia induced by ABT-263 is dose-dependent and reversible effect[64], therefore it is possibility to control thrombocytopenia by appropriate terms of treatment or amounts of BCLX inhibitors[65, 66]. It is also another possibility to develop the drug delivery system to virus-infected cells to avoid the unexpected side effects. Further studies are needed to use inhibitors of BCLX for antiviral therapy. Based on these results, we propose the model as shown in Fig 7 to explain the regulation of BCL2 family proteins upon infection with flaviviruses. In this study, we showed that infection with flaviviruses reduces expression of MCL1 but remaining BCLXL delayed apoptosis in infected cells. This delay of apoptosis is crucial for viral dissemination to neighbouring cells, leading to high pathogenicity. On the other hand, impairment of BCLXL accelerates apoptosis in cells infected with flavivirus and dissemination is reduced by the removal of infected cells through phagocytosis, leading to local infection and low pathogenicity in vivo. In conclusion, we have demonstrated that BCLXL is a critical cell survival factor during infection with flaviviruses, and that inhibition of BCLXL by treatment with BH3 mimetics restricts the production of infectious particles in vitro and in vivo. We showed that expression of pro-survival MCL1 is reduced upon infection with flaviviruses. Although further studies are needed to clarify the molecular mechanisms, the translational suppression induced by viral infection is suggested to participate in the instability of unstable proteins including MCL1. Bclx+/- mice exhibited impairment of pathogenicity to JEV infection, together with the enhanced viral clearance by phagocytosis of infected cells through the stimulation of apoptosis by BCLXL inhibition. In addition, Bclx+/- mice, and wild-type mice that received ABT-263, exhibited prolonged survival rates after JEV challenge compared with untreated wild-type mice. These data suggest that BCLXL may serve as a novel antiviral target that facilitates suppression of the propagation of a broad range of the family Flaviviridae viruses.
The genus Flavivirus including Japanese encephalitis virus , Dengue virus , and Zika virus all of which are mosquito-borne human pathogen and cause serious diseases in humans . Therefore , the development of effective vaccines and antivirals against several flaviviruses is still needed . BCL2 family proteins control mitochondria-mediated apoptosis to maintain cell homeostasis via the removal of damaged cells and pathogen-infected cells , deregulation of which leads to severe diseases including cancer and autoimmune diseases . Here , we showed that BCLXL is a critical cell survival factor during infection with flaviviruses , and that inhibition of BCLXL by treatment with BH3 mimetics restricts the production of infectious particles and the expression of chemokines in vitro and in vivo . Inhibition of BCLXL induces apoptosis in cells infected with flaviviruses and these cells are quickly removed by engulfment of phagocytes , which leads to inhibition of virus dissemination without any inflammatory reaction . Based on these data , BCLXL would appear to be a suitable target for the development of novel antivirals against a broad range of flavivirus infections .