SnRNA processing

Abstract: One of two general pathways of mRNA decay in the yeast Saccharomyces cerevisiae occurs by deadenylation followed by 3'-to-5' degradation of the mRNA body. Previous results have shown that this degradation requires components of the exosome and the Ski2p, Ski3p, and Ski8p proteins, which were originally identified due to their superkiller phenotype. In this work, we demonstrate that deletion of the SKI7 gene, which encodes a putative GTPase, also causes a defect in 3'-to-5' degradation of mRNA. Deletion of SKI7, like deletion of SKI2, SKI3, or SKI8, does not affect various RNA-processing reactions of the exosome. In addition, we show that a mutation in the SKI4 gene also causes a defect in 3'-to-5' mRNA degradation. We show that the SKI4 gene is identical to the CSL4 gene, which encodes a core component of the exosome. Interestingly, the ski4-1 allele contains a point mutation resulting in a mutation in the putative RNA binding domain of the Csl4p protein. This point mutation strongly affects mRNA degradation without affecting exosome function in rRNA or snRNA processing, 5' externally transcribed spacer (ETS) degradation, or viability. In contrast, the csl4-1 allele of the same gene affects rRNA processing but not 3'-to-5' mRNA degradation. We identify csl4-1 as resulting from a partial-loss-of-function mutation in the promoter of the CSL4 gene. These data indicate that the distinct functions of the exosome can be separated genetically and suggest that the RNA binding domain of Csl4p may have a specific function in mRNA degradation.

Abstract: Cellular homeostasis requires transcriptional outputs to be coordinated, and many events post-transcription initiation can dictate the levels and functions of mature transcripts. To systematically identify regulators of inducible gene expression, we performed high-throughput RNAi screening of the Drosophila Metallothionein A (MtnA) promoter. This revealed that the Integrator complex, which has a well-established role in 3' end processing of small nuclear RNAs (snRNAs), attenuates MtnA transcription during copper stress. Integrator complex subunit 11 (IntS11) endonucleolytically cleaves MtnA transcripts, resulting in premature transcription termination and degradation of the nascent RNAs by the RNA exosome, a complex also identified in the screen. Using RNA-seq, we then identified >400 additional Drosophila protein-coding genes whose expression increases upon Integrator depletion. We focused on a subset of these genes and confirmed that Integrator is bound to their 5' ends and negatively regulates their transcription via IntS11 endonuclease activity. Many noncatalytic Integrator subunits, which are largely dispensable for snRNA processing, also have regulatory roles at these protein-coding genes, possibly by controlling Integrator recruitment or RNA polymerase II dynamics. Altogether, our results suggest that attenuation via Integrator cleavage limits production of many full-length mRNAs, allowing precise control of transcription outputs.

Abstract: Jens Lykke-Andersen, Frank Baas, Joseph Gleeson and colleagues report that mutations in the 3′ exonuclease TOE1 cause pontocerebellar hypoplasia type 7. They further show that these mutations result in the accumulation of incompletely processed small nuclear RNAs, leading to severe, early-onset neurodegeneration.