Heritability of alternative splicing in the human genome

Abstract

Alternative pre-mRNA splicing increases proteomic diversity and provides a potential mechanism underlying both phenotypic diversity and susceptibility to genetic disorders in human populations. To investigate the variation in splicing among humans on a genome-wide scale, we use a comprehensive exon-targeted microarray to examine alternative splicing in lymphoblastoid cell lines (LCLs) derived from the CEPH HapMap population. We show the identification of transcripts containing sequence verified exon skipping, intron retention, and cryptic splice site usage that are specific between individuals. A number of novel alternative splicing events with no previous annotations in either the RefSeq and EST databases were identified, indicating that we are able to discover de novo splicing events. Using family-based linkage analysis, we demonstrate Mendelian inheritance and segregation of specific splice isoforms with regulatory haplotypes for three genes: OAS1, CAST, and CRTAP. Allelic association was further used to identify individual SNPs or regulatory haplotype blocks linked to the alternative splicing event, taking advantage of the high-resolution genotype information from the CEPH HapMap population. In one candidate, we identified a regulatory polymorphism that disrupts a 5Ј splice site of an exon in the CAST gene, resulting in its exclusion in the mutant allele. This report illustrates that our approach can detect both annotated and novel alternatively spliced variants, and that such variation among individuals is heritable and genetically controlled.[The microarray data from this study have been submitted to GEO under accession no. GES7952.]The human genome is estimated to contain ∼20,000-25,000 genes, and recent studies suggest that ∼50%-75% of multi-exon genes undergo alternative splicing (AS), generating multiple mRNA isoforms and greatly increasing human proteomic diversity (Lander et al. 2001;Modrek et al. 2001). The splicing of mRNA is a highly regulated process involving the interactions of trans-acting splicing factors and cis-acting regulatory motifs. Disruptions of this process through mutations within these factors and regulatory signals may play an important role in phenotypic diversity and genetic disorders ( ) and applied to identification of AS events that are tissue-specific, for the most part. However, one caveat of these studies utilizing customized arrays is a bias toward genes with solid EST and cDNA evidence for known AS events and that are therefore limited in their usefulness as a discovery tool for de novo splicing events. Here, we have chosen to use an alternative array design, the Affymetrix GeneChip Human Exon 1.0 ST Array, which is less biased toward known AS events by targeting multiple probes to individual exons and allowing simultaneous, exon-level detection of expression levels for 1.4 million probe sets covering over one million known and predicted human exons (Fig. 1). Exon-tiling arrays have several advantages over exon-junction arrays: flexibility of probe placement, exact trans...

Common polymorphic transcript variation in human disease

“…OAS1 is an enzyme involved in innate antiviral response with splice variants that are known to affect susceptibility to viral infection (Bonnevie-Nielsen et al 2005) and multiple sclerosis (Fedetz et al 2006). The same splice site SNP known to associate with disease susceptibility was recently shown also to associate with splicing of OAS1 in CEPH cell lines (Kwan et al 2007), a finding that is confirmed by our analysis (Supplemental Fig. 2; P < 10 À40 ).…”

“…A previous study using the same exon arrays on two HapMap CEPH cell lines (Kwan et al 2007) reported finding 8771 heritable probe sets at a 1% FDR. This is somewhat greater than our figure of 6536 probe sets at a 5.2% FDR.…”

“…1B), suggesting that variation near the 39-ends of transcripts (including alternative 39-UTRs and alternative polyadenylation sites) tends to show the strongest genetic determination. The top two genes (IRF5 and OAS1) (Fig 2A,B; Table 1) have both previously been shown to exhibit genetically controlled PTV in these cell lines (Bonnevie-Nielsen et al 2005;Graham et al 2007;Kwan et al 2007). IRF5 is a transcription factor that acts downstream from Toll-like receptors and has been implicated as The distribution of heritabilities (h 2 ) for the normalized expression of 170,956 probe sets.…”

Systematic evaluation of the effect of common SNPs on pre-mRNA splicing

“…Recent largescale studies have suggested that a relatively high proportion of human genes show variation in expression due to allele-dependent splicing. In the few experiments approaching the last phenomenon from a transcript-based perspective, known SNPs identified as candidates for a splicing effect were tested with isoform-specific PCR (Hull et al, 2007), or chip-based methods were used to seek evidence for alternative splicing (Kwan et al, 2007;Kwan et al, 2008). Hull et al (2007) analyzed the splicing patterns of 250 exons in 22 individuals who had been previously genotyped as part of the HapMap project.…”

Using positional distribution to identify splicing elements and predict pre-mRNA processing defects in human genes

“…Transcript analysis of genotyped cell lines has discovered numerous cases of allelic splicing demonstrating that polymorphisms also disrupt splicing (11,12). These types of functional variants likely account for a similarly large fraction of the detected genetic risk for complex disease and could eventually be a target for molecular intervention.…”