Dataset Information

Identification of allele-specific alternative mRNA processing via transcriptome sequencing.

ABSTRACT: Establishing the functional roles of genetic variants remains a significant challenge in the post-genomic era. Here, we present a method, allele-specific alternative mRNA processing (ASARP), to identify genetically influenced mRNA processing events using transcriptome sequencing (RNA-Seq) data. The method examines RNA-Seq data at both single-nucleotide and whole-gene/isoform levels to identify allele-specific expression (ASE) and existence of allele-specific regulation of mRNA processing. We applied the methods to data obtained from the human glioblastoma cell line U87MG and primary breast cancer tissues and found that 26-45% of all genes with sufficient read coverage demonstrated ASE, with significant overlap between the two cell types. Our methods predicted potential mechanisms underlying ASE due to regulations affecting either whole-gene-level expression or alternative mRNA processing, including alternative splicing, alternative polyadenylation and alternative transcriptional initiation. Allele-specific alternative splicing and alternative polyadenylation may explain ASE in hundreds of genes in each cell type. Reporter studies following these predictions identified the causal single nucleotide variants (SNVs) for several allele-specific alternative splicing events. Finally, many genes identified in our study were also reported as disease/phenotype-associated genes in genome-wide association studies. Future applications of our approach may provide ample insights for a better understanding of the genetic basis of gene regulation underlying phenotypic diversity and disease mechanisms.

SUBMITTER: Li G

PROVIDER: S-EPMC3401465 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

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Identification of allele-specific alternative mRNA processing via transcriptome sequencing.

Li Gang G Bahn Jae Hoon JH Lee Jae-Hyung JH Peng Guangdun G Chen Zugen Z Nelson Stanley F SF Xiao Xinshu X

Nucleic acids research 20120329 13

Establishing the functional roles of genetic variants remains a significant challenge in the post-genomic era. Here, we present a method, allele-specific alternative mRNA processing (ASARP), to identify genetically influenced mRNA processing events using transcriptome sequencing (RNA-Seq) data. The method examines RNA-Seq data at both single-nucleotide and whole-gene/isoform levels to identify allele-specific expression (ASE) and existence of allele-specific regulation of mRNA processing. We app ...[more]

PMID: 22467206

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Project description:Splice isoform structure and abundance can be affected by either noncoding or masquerading coding variants that alter the structure or abundance of transcripts. When these variants are common in the population, these nonconstitutive transcripts are sufficiently frequent so as to resemble naturally occurring, alternative mRNA splicing. Prediction of the effects of such variants has been shown to be accurate using information theory-based methods. Single nucleotide polymorphisms (SNPs) predicted to significantly alter natural and/or cryptic splice site strength were shown to affect gene expression. Splicing changes for known SNP genotypes were confirmed in HapMap lymphoblastoid cell lines with gene expression microarrays and custom designed q-RT-PCR or TaqMan assays. The majority of these SNPs (15 of 22) as well as an independent set of 24 variants were then subjected to RNAseq analysis using the ValidSpliceMut web beacon (http://validsplicemut.cytognomix.com), which is based on data from the Cancer Genome Atlas and International Cancer Genome Consortium. SNPs from different genes analyzed with gene expression microarray and q-RT-PCR exhibited significant changes in affected splice site use. Thirteen SNPs directly affected exon inclusion and 10 altered cryptic site use. Homozygous SNP genotypes resulting in stronger splice sites exhibited higher levels of processed mRNA than alleles associated with weaker sites. Four SNPs exhibited variable expression among individuals with the same genotypes, masking statistically significant expression differences between alleles. Genome-wide information theory and expression analyses (RNAseq) in tumor exomes and genomes confirmed splicing effects for 7 of the HapMap SNP and 14 SNPs identified from tumor genomes. q-RT-PCR resolved rare splice isoforms with read abundance too low for statistical significance in ValidSpliceMut. Nevertheless, the web-beacon provides evidence of unanticipated splicing outcomes, for example, intron retention due to compromised recognition of constitutive splice sites. Thus, ValidSpliceMut and q-RT-PCR represent complementary resources for identification of allele-specific, alternative splicing.

Dataset Information

Identification of allele-specific alternative mRNA processing via transcriptome sequencing.

Publications

Identification of allele-specific alternative mRNA processing via transcriptome sequencing.

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