UCLA

Because RNA-synthesis and RNA-processing are spatially and temporally coordinated, RNA splicing takes place in the context of chromatin. H2A.Z is a highly conserved histone variant of the canonical histone H2A. In Saccharomyces cerevisiae, the SWR-C complex deposits H2A.Z into chromatin near the beginning of protein-coding genes where it helps regulate transcription. In this dissertation we elucidate a role for H2A.Z in coordinating the processes of RNA transcription and pre-mRNA splicing.

H2A.Z is required for optimal splicing of intron-containing genes, particularly under suboptimal splicing conditions. H2A.Z genetically interacts with the, particularly with U2 snRNP complex, and is required for efficient spliceosome rearrangements. Loss of H2A.Z results in defective spliceosome rearrangements, particularly those involving the U2 snRNP. H2A.Z loss impairs transcription elongation, suggesting that spliceosome rearrangements are tied to the role of H2A.Z in elongation. Depletion of disassembly factor Prp43 suppresses H2A.Z-mediated splice defects, indicating that, in the absence of H2A.Z, stalled spliceosomes are disassembled and unspliced RNAs are released. These data demonstrate that H2A.Z is required for efficient pre-mRNA splicing and indicate a role for H2A.Z in coordinating the kinetics of transcription elongation and splicing.

Chromatin not only affects splicing locally but also globally. We demonstrate that a chromatin remodeler regulates respiration through modulation spliceosome availability. Nutrient-responsive decrease in the Snf2 chromatin remodeler leads to ribosomal protein gene (RPG) down-regulation. Because RPGs are intron-enriched and highly transcribed, this relieves competition for limiting spliceosomes and allows for increased splicing of weaker substrates, such as PTC7. The spliced PTC7 isoform encodes a mitochondrial protein that promotes Coenzyme Q6 biosynthesis during respiration. These findings establish a role for the SWI/SNF complex in yeast in transition to respiratory metabolism through global regulation of splicing.

Furthermore, we show that disruption of transcription elongation kinetics can effect splicing both locally and globally, resulting in multi-faceted changes in splicing outcomes. While loss of the elongation factor Dst1 exacerbates transcription and splicing defects in cells lacking H2A.Z, it can also improve splicing of many intron-containing genes. In the absence of both Dst1 and H2A.Z, RPGs are down-regulated, allowing for spliceosome redistribution to and increased splicing of weaker substrates.

Additionally, we show that H2A.Z is required for growth in alkaline pH and expression of phosphate-starvation response genes. However, we find that, when H2A.Z is lost, deletion of Swr1 suppresses alkaline sensitivity, relieves aberrant RNA polymerase II (RNAPII) phosphorylation, and restores appropriate phosphate-gene. Therefore, stress sensitivity and transcription misregulation may be due to deleterious effects of an incomplete SWR-C complex, demonstrating the need for precise control of chromatin remodeling activity in the coordination of gene expression processes.