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: Synergistic assembly of human pre-spliceosomes across introns and exons}
Article:
Abstract:
Most human genes contain multiple introns, necessitating mechanisms to effectively define exons and ensure their proper connection by spliceosomes. Human spliceosome assembly involves both cross-intron and cross-exon interactions, but how these work together is unclear. We examined in human nuclear extracts dynamic interactions of single pre-mRNA molecules with individual fluorescently tagged spliceosomal subcomplexes to investigate how cross-intron and cross-exon processes jointly promote pre-spliceosome assembly. U1 subcomplex bound to the 5' splice site of an intron acts jointly with U1 bound to the 5' splice site of the next intron to dramatically increase the rate and efficiency by which U2 subcomplex is recruited to the branch site/3' splice site of the upstream intron. The flanking 5' splice sites have greater than additive effects implying distinct mechanisms facilitating U2 recruitment. This synergy of 5' splice sites across introns and exons is likely important in promoting correct and efficient splicing of multi-intron pre-mRNAs.
Introduction:
Spliceosomes consist of the U1, U2, and U4/U6. U5 small nuclear ribonucleoproteins( snRNPs) and multiprotein Prp19-complex as major building blocks, plus many transiently interacting splicing factors( Wahl et al., 2009). This machinery recognizes and assembles stepwise at splice sites( SS)( U1 at 5'SS and U2 at the 3'SS/branch site) to form pre-spliceosomes, which are subsequently remodeled into catalytically active spliceosomes. Pre-spliceosomes can form on multi-intron pre-mRNAs through at least two different pathways. An intron can be recognized a) via cross-intron interactions leading directly to a catalytically active spliceosome, or b) via cross-exon interactions where the exons flanking an intron are first defined, after which cross-intron interactions between adjacent cross-exon complexes lead to spliceosome assembly( Moldón and Query, 2010). On human pre-mRNAs, which characteristically harbor multiple long introns and short exons, exon definition predominates( Berget, 1995; Fox-Walsh et al., 2005). Indeed, splicing is greatly enhanced when a 5'SS is present across the exon downstream of an intron, highlighting the importance of exon definition in humans( Talerico and Berget, 1990; Yue and Akusjärvi, 1999). Cross-exon pre-spliceosomes can transition into cross-intron pre-spliceosomes, each having a distinct protein composition; the latter can then productively splice the pre-mRNA( Chiara and Reed, 1995; Schneider et al., 2010). However, the mechanisms by which cross-intron and cross-exon pre-spliceosomes work together to facilitate pre-mRNA splicing remain unclear. In S. cerevisiae, where the cross-intron pathway predominates, single-molecule approaches have proven invaluable for elucidating the kinetic pathways and subcomplex dynamics involved in spliceosome assembly( Semlow et al., 2016; Warnasooriya and Rueda, 2014). Here we developed the tools necessary to implement colocalization single-molecule spectroscopy( CoSMoS) in human cell extracts and used this system to investigate the dynamic mechanism of cross-intron and cross-exon cooperation in human pre-spliceosome assembly.
Discussion:
Here we establish methods for observing the dynamics of spliceosomal subcomplexes on and the splicing of individual pre-mRNA molecules in human cell extract. The experiments reveal that the dynamics of U1 binding to the 5'SS are largely unaffected by the presence of U2 binding sites positioned either downstream( cross-exon) or upstream( cross-intron) of the 5'SS. In contrast, stable U2 binding to the 3'SS is accelerated by a 5'SS situated across either the adjacent intron or adjacent exon. Strikingly, when functional 5'SS are present together at both cross-intron and cross-exon locations, they synergistically promote U2 recruitment. The more than additive effect of the flanking 5'SS indicates that U1 employs different molecular mechanisms/interactions across introns and across exons to accelerate U2 recruitment. A simple model that would explain this observed synergy of cross-exon and cross-intron 5'SS on U2 recruitment rate is that cross-exon U1 binding assists in recruiting the splicing factors that form a platform for U2 binding to the upstream 3'SS region, whereas the cross-intron U1-U2 interaction favors a U2 conformation capable of productive branch site engagement( Figure 5, left). Across exons, U1 is known to recruit U2AF( U2 auxiliary factor), which recognizes the polypyrimidine tract and 3'SS AG, and physically interacts with other proteins( e. g., SF1, p14, SF3B and SR proteins) required for stable U2 binding( Black, 2003). Across introns, U1 and U2 can interact via the DEAD-box protein Prp5, the ATPase activity of which promotes a structural change in U2 that makes the branch site recognition sequence more solvent accessible( Abu Dayyeh et al., 2002; O'Day et al., 1996; Xu et al., 2004). A different and not necessarily mutually exclusive model is that cross-exon and cross-intron U1 interactions could both help form the binding platform for U2, but by interacting with different proteins( e. g., the upstream U1 stabilizes SF1 and the downstream U1 stabilizes U2AF)( Black, 2003). This arrangement would increase the likelihood of U2 encountering a fully assembled binding platform when in a conformation capable of stable branch site engagement( Figure 5, right). Other mechanisms for synergy, such as those mediated by effects on RNA secondary structure, are also possible. Models of the type presented in Figure 5 assume that once a functional U1-U2 pre-spliceosome forms across the intron, interactions with U1 bound to the downstream 5'SS provide no additional U2 binding stabilization or stimulation of subsequent steps of spliceosome assembly( Figure 5, bottom). Such models predict that the presence of a downstream U1 will not affect the lifetimes of stable U2 complexes once formed. The data in Figure 4C are consistent with this prediction: both the overall shape of dwell time distributions and the ratios of binding frequency to occupancy are similar for the RNAs with( 5i3e5: 1. 0 ± 0. 2 s−1) or without( 5i3eX: 1. 3 ± 0. 9 s−1) the downstream site. Taken together, our data suggest that the synergistic stimulatory effect of the downstream 5'SS on splicing is exerted at the U2 recruitment step and the processes that enable it, not at the subsequent steps of spliceosome assembly and splicing. Inappropriate skipping of otherwise constitutive internal exons can occur with exceptionally low frequency( ~1 in 105 splicing events)( Fox-Walsh and Hertel, 2009). But the molecular mechanisms contributing to this remarkable accuracy were previously unclear. By implementing CoSMoS in extracts from human cells expressing genetically-tagged proteins, we here show that a major contributor to exon inclusion is collaboration between flanking 5'SS. This collaboration dramatically increases the rate of stable U2 recruitment during pre-spliceosome formation. Cross-intron and cross-exon 5'SS synergy on U2 recruitment rate explains how U1 can have such a strong effect on U2 recruitment despite its association with pre-mRNA being much more dynamic than U2. This mechanism is likely crucial for rapid definition of internal exons in multi-intron RNAs, enabling the human splicing machinery to avoid inappropriate exon skipping.
A gene is a segment of DNA that usually carries the information required to build a protein , the molecules responsible for most of life’s processes . This DNA segment is organized in modules , with coding sections separated by portions of non-coding DNA known as introns . When a gene is ‘turned on’ , it gets faithfully copied into a molecule of pre-messenger RNA ( pre-mRNA ) , which contains the alternating coding and non-coding modules . Before it can serve as a template to create a protein , this pre-mRNA must be processed and all the introns removed by a structure called the spliceosome . If this delicate process goes wrong , inaccurate protein templates are produced that may be damaging for the cell . Spliceosomes are precise molecular ‘scissors’ that can recognize where a coding module stops and an intron starts , and then make a snip in the pre-mRNA to remove the non-coding sequence . The spliceosome is a complex molecular machine formed of numerous parts – including one known as U1 snRNP – that must come together . When a pre-mRNA has several introns , a spliceosome assembles anew for each of them . Braun et al . designed a new method that allows them to ‘tag’ spliceosomes extracted from a human cell and follow them as they come together . The experiments show that spliceosomes working on different introns in the same pre-mRNA actually help each other out . As one assembles , this helps the spliceosome that processes the neighboring intron to get built . In particular , the U1 snRNPs processing nearby introns collaborate to promote the assembly and activity of the spliceosomes . This teamwork is likely important to guarantee that multiple introns are cut out quickly and accurately .