UC Santa Cruz

The spliceosome, a large macromolecular machine that removes introns from pre-mRNA to form mature mRNA, contains five small RNAs that play key roles. To learn more about the role played by one of these, U2, we started with a temperature-sensitive mutant of U2, U33Δ. Using U33Δ, we analyzed secondary mutations that restored growth at a non-permissive temperature in Saccharomyces cerevisiae. We found that this suppressor mutation was in a gene, NOB1, that is unrelated to splicing and instead is a crucial component of ribosomal RNA processing. This mutation, found to be a delta (δ) insert from a Ty transposable element, causes reduced function in Nob1 and an accumulation of unprocessed rRNA. RNA sequencing revealed U33Δ has a splicing defect and change in gene expression, both of which the nob1-Tyδ allele restored. How this rRNA processing defect rescues a spliceosome mutant led us to investigate this phenomenon through the lens of systems biology, and we found that decreasing the load of pre-mRNAs or increasing the capacity of the spliceosome were both able to restore the growth of U33Δ. We found new mechanisms that can change the pre-mRNA pool that rescue mutations in the spliceosome, adding evidence to the hypothesis that splicing regulation is directly related to the spliceosome’s capacity and workload.