| Efficient termination of transcription plays an important role in cell homeostasis by preventing spurious transcription that might interfere with cis-acting regulatory elements found downstream of genes, by ensuring synthesis of functional transcripts, and by promoting recycling of factors needed for a new transcription cycle.;Another technique was high-copy suppression of growth and termination defects of mutants encoding factors required for snoRNA termination leading to the identification of Sen1, which, by re-anchoring the yeast type I protein phosphatase to the machinery required for snoRNA termination, restored a dephosphorylation-dependent event regulating this process.;Finally, systematic screening of mutants for termination defects of a reporter construct also identified new factors required for termination of mRNAs, including Sen1. Genetic and biochemical techniques were used to support a general role in mRNA termination for Sen1 and proved that termination of different classes of transcripts synthesized by RNAPII occurs through a common mechanism.;I used both biochemical and genetic approaches to identify new factors required for termination by RNAPII. One was isolation by tandem affinity purification of complexes containing known and putative termination factors and identification by mass spectrometry of the specifically associated proteins. Systematic purification of these associated proteins allowed identification of the core components of protein complexes, as well as less stable bridges between complexes. In addition, purification of complexes from strains in which a component of the complex was absent or mutated permitted identification of factors playing structural roles within the complex, as well as interactions between the components of the complex. |
