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Ribosomal footprinting and RNASeq in two strains of yeast and their diploid hybrid

ABSTRACT: Heritable differences in gene expression between individuals are an important source of phenotypic variation. The question of how closely the effects of genetic variation on protein levels mirror those on mRNA levels remains open. Here, we addressed this question by using ribosomal footprinting to examine how genetic differences between two strains of the yeast S. cerevisiae affect translation. Strain differences in translation were observed for hundreds of genes, more than half as many as showed genetic differences in mRNA levels. Similarly, allele specific measurements in the diploid hybrid between the two strains found roughly half as many cis-acting effects on translation as were observed for mRNA levels. In both the parents and the hybrid, strong effects on translation were rare, such that the direction of an mRNA difference was typically reflected in a concordant footprint difference. The relative importance of cis and trans acting variation on footprint levels was similar to that for mRNA levels. Across all expressed genes, there was a tendency for translation to more often reinforce than buffer mRNA differences, resulting in footprint differences with greater magnitudes than the mRNA differences. Finally, we catalogued instances of premature translation termination in the two yeast strains. Overall, genetic variation clearly influences translation, but primarily does so by subtly modulating differences in mRNA levels. Translation does not appear to create strong discrepancies between genetic influences on mRNA and protein levels.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE55400 | GEO | 2014/03/31

SECONDARY ACCESSION(S): PRJNA239467

REPOSITORIES: GEO

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Project description:Gene expression levels are determined by the balance between rates of mRNA transcription and decay, and genetic variation in either of these processes can result in heritable differences in transcript abundance. Although the genetics of gene expression has been the subject of intense interest, the contribution of heritable variation in mRNA decay rates to gene expression variation has received far less attention. To this end, we developed a novel statistical framework and measured allele-specific differences in mRNA decay rates in a diploid yeast hybrid created by mating two genetically diverse parental strains. In total, we estimate that 31% of genes exhibit allelic differences in mRNA decay rate, of which 350 can be identified at a false discovery rate of 10%. Genes with significant allele-specific differences in mRNA decay rate have higher levels of polymorphism compared to other genes, with all gene regions contributing to allelic differences in mRNA decay rate. Strikingly, we find widespread evidence for compensatory evolution, such that variants influencing transcriptional initiation and decay having opposite effects, suggesting steady-state gene expression levels are subject to pervasive stabilizing selection. Our results demonstrate that heritable differences in mRNA decay rates are widespread, and are an important target for natural selection to maintain or fine-tune steady-state gene expression levels. We measured rates of allele-specific mRNA decay (ASD) in a diploid yeast produced by mating two genetically diverse haploid Saccharomyces cerevisiae strains: the laboratory strain BY4716 (BY), which is isogenic to the reference sequence strain S288C, and the wild Californian vineyard strain RM11-1a (RM). Briefly, we introduced rpb1-1, a temperature sensitive mutation in an RNA polymerase II subunit, to each of the haploid yeast strains, mated the strains, and grew the resulting hybrid diploid to mid-log phase at 24 M-BM-0C, before rapidly shifting the culture to 37 M-BM-0C to inhibit transcription. RNA-seq was performed on culture samples taken at 0, 6, 12, 18, 24, and 42 minutes subsequent to the temperature shift. To identify ASD, we used transcribed polymorphisms to distinguish between parental transcripts, and compared the relative levels of transcript abundance over the time course. Note, this experimental design internally controls for trans-acting regulatory variation as well as environmental factors. Under the null hypothesis of no ASD, the proportion of reads from the BY transcript (p_BY = N_BY / (N_BY + N_RM)) observed over the time course remains unchanged. However, genes with ASD will exhibit an increasing or decreasing proportion of BY reads as a function of time. In total, we measured ASD from three independent biological replicates.

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Ribosomal footprinting and RNASeq in two strains of yeast and their diploid hybrid

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