Heritable variation of mRNA decay rates in yeast.

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 a 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. We estimate that 31% of genes exhibit allelic differences in mRNA decay rates, of which 350 can be identified at a false discovery rate of 10%. Genes with significant allele-specific differences in mRNA decay rates have higher levels of polymorphism compared to other genes, with all gene regions contributing to allelic differences in mRNA decay rates. Strikingly, we find widespread evidence for compensatory evolution, such that variants influencing transcriptional initiation and decay have opposite effects, suggesting that 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.

Massively parallel functional dissection of mammalian enhancers in vivo.

The functional consequences of genetic variation in mammalian regulatory elements are poorly understood. We report the in vivo dissection of three mammalian enhancers at single-nucleotide resolution through a massively parallel reporter assay. For each enhancer, we synthesized a library of >100,000 mutant haplotypes with 2-3% divergence from the wild-type sequence. Each haplotype was linked to a unique sequence tag embedded within a transcriptional cassette. We introduced each enhancer library into mouse liver and measured the relative activities of individual haplotypes en masse by sequencing the transcribed tags. Linear regression analysis yielded highly reproducible estimates of the effect of every possible single-nucleotide change on enhancer activity. The functional consequence of most mutations was modest, with ∼22% affecting activity by >1.2-fold and ∼3% by >2-fold. Several, but not all, positions with higher effects showed evidence for purifying selection, or co-localized with known liver-associated transcription factor binding sites, demonstrating the value of empirical high-resolution functional analysis.