Integration Of Splicing Regulation With Gene Expression Programs

Genes expressed within a cell define its identity and functional state. Gene expression in eukaryotes is highly regulated from transcription, to RNA processing, to translation. Pre-mRNA splicing, or the removal of intervening intronic sequences and ligation of expressed exonic sequences, is an essential step of RNA processing and contributes to regulation of the quantity and coding potential of the mRNA. Splicing can vary in complexity from the simpler ligation of neighboring exons, to more complex patterns of alternative splicing where exons or parts of exons may be differentially included or skipped. In order to understand how splicing regulation contributes to gene expression programs as a whole, splicing must be integrated with transcriptional networks and post-transcriptional networks that work together to drive transitions through cell states. The studies presented here address how splicing regulation is integrated with the meiotic developmental gene expression program in budding yeast. Meiosis is known to depend on both transcription and splicing regulation. In particular, the splicing activator Mer1 is required for meiosis. The genome-wide studies in Chapter 2 define the entire Mer1 splicing network to consist of Mer1 and four regulated transcripts. The contribution of Mer1 to the meiotic gene expression program is also explored. These studies indicate that transcription of the Mer1 network is activated by the first transcriptional wave of meiosis, which is driven by the transcriptional regulators Ume6 and Ime1. We show that co-induction of Mer1 with its regulated transcripts creates a delay in expression of the Mer1-regulated transcripts that depends on accumulation of Mer1 protein. Subsequently expression of two Mer1-target transcripts is required for induction of the second transcriptional wave in meiosis, activated by the Ndt80 transcription factor, and progression through the meiotic program. Thus, the Mer1 splicing network links the early Ume6-dependent transcriptional wave with the second Ndt80-dependent transcriptional wave in meiosis. This study reveals how splicing networks may be interlaced with transcriptional networks to drive progression through a gene expression program. Genome-wide analyses of splicing during meiosis presented in Chapter 2 also revealed a general increase in splicing efficiency. The global splicing improvement is coincident with the transcriptional repression of ribosomal protein genes (RPGs), which constitute a majority of intron-containing transcripts in the cell. The studies presented in Chapter 3 identify the molecular mechanism for the splicing improvement during meiosis as a relief in competition between pre-mRNAs for the spliceosome. Although relief in competition between transcripts and improved splicing of non-RPG pre-mRNAs is hardwired into the meiotic gene expression program, vegetative cells where RPGs are transcriptional repressed also display improved splicing of other transcripts. This study is the first to show that global splicing regulation depends on the effective load of pre-mRNAs on the splicing machinery. This regulatory mechanism that we are the first to describe is called trans-competition control. The studies presented in this thesis contribute to understanding how splicing regulation is coordinately integrated with transcriptional networks to promote progression through a gene expression program.