Regulation of alternative splicing by the core spliceosomal machinery

Abstract

Alternative splicing (AS) plays a major role in the generation of proteomic diversity and in gene regulation. However, the role of the basal splicing machinery in regulating AS remains poorly understood. Here we show that the core snRNP (small nuclear ribonucleoprotein) protein SmB/B9 self-regulates its expression by promoting the inclusion of a highly conserved alternative exon in its own pre-mRNA that targets the spliced transcript for nonsense-mediated mRNA decay (NMD). Depletion of SmB/B9 in human cells results in reduced levels of snRNPs and a striking reduction in the inclusion levels of hundreds of additional alternative exons, with comparatively few effects on constitutive exon splicing levels. The affected alternative exons are enriched in genes encoding RNA processing and other RNA-binding factors, and a subset of these exons also regulate gene expression by activating NMD. Our results thus demonstrate a role for the core spliceosomal machinery in controlling an exon network that appears to modulate the levels of many RNA processing factors.[Keywords: alternative splicing; Sm proteins; snRNP; autoregulation; NMD; exon network] Supplemental material is available for this article. The production of multiple mRNA variants through alternative splicing (AS) is estimated to take place in transcripts from >95% of human multiexon genes Wang et al. 2008). AS represents a mechanism for gene regulation and expansion of the proteome. The most widely studied trans-acting factors regulating AS are proteins of the SR (Ser/Arg-rich) and hnRNP (heterogeneous ribonucleoprotein) families, as well as numerous tissue-restricted AS factors (for review, see Chen and Manley 2009;Nilsen and Graveley 2010). These factors generally regulate AS by recognizing cis-acting sequences in exons or introns and by promoting or suppressing the assembly of the spliceosome at adjacent splice sites. In contrast to these AS regulatory factors, much less is known about how or the extent to which components of the basal or ''core'' splicing machinery modulate splice site decisions.The spliceosome is a large RNP complex that carries out the removal of introns from pre-mRNAs. It comprises the U1, U2, U4/6, and U5 small nuclear RNPs (snRNPs) and several hundred protein factors (for review, see Wahl et al. 2009). Studies in yeast and metazoan systems have indicated that the levels of some of these core splicing components can affect splice site choice. Microarray profiling revealed transcript-specific effects on splicing in yeast strains harboring mutations in or deletions of core splicing components Pleiss et al. 2007). Knockdown of several core splicing factors in Drosophila cells resulted in transcript-specific effects on AS reporters (Park et al. 2004). Deficiency of the snRNP assembly factor SMN (survival of motor neuron) in a mouse model of spinal muscular atrophy (SMA) resulted in tissue-specific perturbations in snRNP levels and splicing defects (Gabanella et al. 2007;Zhang et al. 2008). Tiling microarray profiling analysis of fission yeast...

The spliceosome assembly factor GEMIN2 attenuates the effects of temperature on alternative splicing and circadian rhythms

“…We observed that GEMIN2 had a greater effect on alternative than on constitutive splicing, a phenomenon that has already been reported for several core snRNP components, such as SmB (19), LSm4 (20), or U1C (21) (Fig. 2B).…”

Role for LSM genes in the regulation of circadian rhythms

“…Indeed, core components of the promoter recognition complex, generally considered to have a passive role in controlling gene expression, were recently shown to regulate cell-specific transcriptional programs during development (38). Similarly, down-regulation of SmB, a core component of U1-U5 snRNPs, in human cells, or mutations in the zebrafish U1C gene, which encodes a component of U1SnRNP, result in stronger alterations in AS than in constitutive splicing (21)(22)(23). A genomewide characterization of RNA processing reported recently for sad1/lsm5 also revealed alterations in a subset of splicing events (29).…”

“…GSE60387). [21][22][23]. Thus, changes in the levels and/or activities of corespliceosomal proteins may play a regulatory role in AS, although specific signaling pathways where this is relevant are not known.…”

Introns in UTRs: Why we should stop ignoring them

“…The most dramatic functional enrichment among NMD-affected transcripts is for those encoding RNA-binding proteins [45,66]. Within this class of genes, exons that introduce stop codons upstream of introns are extremely wellconserved [44,67].…”

“…That is, as an RNA binding protein increases in abundance, it increasingly binds its own pre-mRNA and facilitates production of a 3UI-containing form subject to NMD, thereby maintaining protein homeostasis [44,67,68]. Proteins undergoing this type of regulation include ribosomal proteins [42,[69][70][71], core spliceosomal proteins [66,72] and alternative splicing regulators such as hnRNP and SR proteins [45,67]. In fact, every one of the 11 human SR proteins has a 3UI isoform and regulates its own production by AS-NMD [67].…”