Abstract
mRNA synthesis, processing, and destruction involve a complex series of molecular steps that are incompletely understood. Because the RNA intermediates in each of these steps have finite lifetimes, extensive mechanistic and dynamical information is encoded in total cellular RNA. Here we report the development of SnapShot-Seq, a set of computational methods that allow the determination of in vivo rates of pre-mRNA synthesis, splicing, intron degradation, and mRNA decay from a single RNA-Seq snapshot of total cellular RNA. SnapShot-Seq can detect in vivo changes in the rates of specific steps of splicing, and it provides genome-wide estimates of pre-mRNA synthesis rates comparable to those obtained via labeling of newly synthesized RNA. We used SnapShot-Seq to investigate the origins of the intrinsic bimodality of metazoan gene expression levels, and our results suggest that this bimodality is partly due to spillover of transcriptional activation from highly expressed genes to their poorly expressed neighbors. SnapShot-Seq dramatically expands the information obtainable from a standard RNA-Seq experiment.
Highlights
The expression level of an individual mRNA species depends on the rates of key events at three phases of its lifecycle: pre-mRNA transcription, pre-mRNA processing, and mRNA degradation [1]
We describe a new method – SnapShot-Seq – for quantifying mammalian mRNA dynamics, using only standard RNA sequencing (RNA-Seq) data that can be generated from any total cellular RNA sample
We introduce a quantitative model that relates the densities of total RNA sequencing (RNA-Seq) reads across exons, introns, and splice junctions to the lifetimes of pre-mRNA intermediate species and mature mRNAs
Summary
The expression level of an individual mRNA species depends on the rates of key events at three phases of its lifecycle: pre-mRNA transcription, pre-mRNA processing, and mRNA degradation [1]. Each of these steps has the potential to be regulated to control gene expression. The estimated time required for splicing ranges from 30 seconds [8]to 12 minutes [9] in studies of individual introns [8,9,10,11,12] _ENREF_10 It is unclear whether this variability reflects intronto-intron variation, species-to-species variation, or the differences in the methods used. The ability to assess in vivo splicing rates genome-wide could reveal new modes of gene regulation and identify functions for the many putative splicing factors whose functions remain unknown
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