MRC Human Genetics Unit, Edinburgh, EH4 2XU, UK
(e-mail: jeremy.sanford{at}hgu.mrc.ac.uk; javier.caceres{at}hgu.mrc.ac.uk)
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Introduction |
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In the nucleus, the C-terminal domain (CTD) of the RNA polymerase II (Pol II) large subunit coordinates many RNA processing events by providing a platform for factors involved in different steps of RNA processing (reviewed by Maniatis and Reed, 2002). Splicing of pre-mRNAs can occur co-transcriptionally (for a review, see Neugebauer, 2002
; Beyer and Osheim, 1988
), and splicing factors that are enriched in interchromatin granule clusters (IGCs) are recruited to the sites of active transcription (Misteli et al., 1997
; Lamond and Spector, 2003
). Components of the capping and polyadenylation machinery also associate with the CTD and these interactions facilitate 5' end cap formation and polyadenylation in vivo (Maniatis and Reed, 2002
). Here, we focus on the central role of pre-mRNA splicing in coordinating many different steps of the gene expression cascade.
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The spliceosome |
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Spliceosome assembly follows a carefully orchestrated stepwise pathway (reviewed by Will and Luhrmann, 2001). This is initiated by recognition of the 5' and 3' splice sites by the U1 snRNP and the heterodimeric U2 snRNP auxiliary factor (U2AF), respectively, generating the E (early) complex. The pre-spliceosomal A complex is generated by the recruitment of the U2 snRNP to the branch point adenosine (BP), in an ATP-dependent manner. Subsequently the U4-U6-U5 tri-snRNP joins the pre-spliceosome to form the B complex, which is resolved to the catalytic C complex following a series of RNA-RNA and RNA-protein rearrangements at the heart of the spliceosome, resulting in the release of U1 and U4. Although the catalytic centre of the spliceosome has not been fully defined, current evidence strongly suggests that pre-mRNA splicing is accomplished by protein-assisted RNA catalysis (reviewed by Nilsen, 2000
).
Many components of this large and complex macromolecular machine have now been identified by proteomic approaches. Indeed, recent studies have led to the identification of over 300 putative spliceosomal protein components (reviewed by Jurica and Moore, 2003); however, many unexpected proteins present, with no apparent direct connection to splicing, still await functional characterization. Some of the additional proteins identified in these complexes have known associations with other aspects of RNA processing, including transcription and mRNA export. This is consistent with the intimate coupling of different steps in gene expression.
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Alternative splicing |
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The highly conserved serine- and arginine-rich (SR) protein family plays an important role in this process (Graveley, 2000). The SR proteins have a modular structure consisting of one or two copies of an RNA-recognition motif (RRM) that determines their RNA-binding specificity, followed by a C-terminal domain rich in alternating serine and arginine residues (the RS domain). SR proteins bound to exonic splicing enhancers (ESEs) promote recruitment of U2AF to the polypyrimidine tract (poly Y) and activate an adjacent 3' splice site. Alternatively, they might facilitate splicing by recruitment of coactivators or antagonize the negative activity of hnRNP proteins recognizing exonic splicing silencer (ESS) elements (reviewed by Blencowe, 2000
; Hastings and Krainer, 2001
). Differences in the activities or amounts of general splicing factors and/or gene-specific splicing regulators during development or in different tissues are thought to cause differential patterns of splicing. Moreover, signal transduction pathways can modulate alternative splice site selection in vivo by regulating the concentration, activity and/or subcellular localization of splicing regulatory proteins (reviewed by Stamm, 2002
).
Another mechanism of regulation of alternative splicing is imparted by the transcriptional machinery and involves the processivity of Pol II (Roberts et al., 1998; de la Mata et al., 2003
). For instance, a slow Pol II, and/or the presence of internal transcriptional pause sites, results in inclusion of the alternative exon harbouring a weak 3' splice site. By contrast, when the same pre-mRNA is transcribed by a highly processive Pol II, the weak alternative 3' splice site is unable to compete with the stronger downstream 3'splice site, which results in skipping of the alternative exon.
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Pre-mRNA splicing and human disease |
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Life after splicing |
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Cytoplasmic activities |
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Acknowledgments |
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References |
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