Coupling between Transcription & Splicing Tunes Gene Expression

Abstract

RNA is synthesized by RNA polymerases that travel along template DNA. The growing RNA polymers are referred to as nascent. As transcription proceeds, nascent RNA grows in length and sequence complexity, increasing opportunity for folding and interacting with RNA binding proteins. In eukaryotes, the capping enzymes modify the mRNA 5’ end, the spliceosome assembles and removes introns, and the polyadenylation machinery cleaves 3’ ends to initiate polyadenylation of mRNA and terminate transcription. All of this takes place in a nascent RNP, which lies close to the DNA axis. I will illustrate how recent single molecule RNA-Seq strategies have revealed the in vivo kinetics of pre-mRNA splicing relative to the progress of RNA polymerase II (Pol II). In budding and fission yeast, the spliceosome can act on introns as soon as they emerge from Pol II, showing that splicing and transcription rates are matched. This predicts that introns present in multi-intron transcripts may be removed in the order of their transcription. Instead, sequencing of full-length nascent RNA molecules in S. pombe revealed that intron removal is equally likely to occur “not in order”, showing regulation. Surprisingly, partially spliced transcripts were rare; most nascent RNA detected was either fully spliced or unspliced, indicating that splicing of any given intron may depend on the splicing status of the other introns in the transcript. Fully unspliced transcripts failed to cleave at the polyA site, underwent transcriptional read-through at gene 3’ends, and were degraded by the exosome. These observations suggest crosstalk among spliceosomes and 3’end processing machineries assembling on the same nascent transcript. I will discuss an example of regulated gene expression in which reduced splicing leads to lower reduced mRNA levels. These findings highlight coordination between transcription and RNA processing steps along the pathway of gene expression.