Browsing by Author "Firestone, Kyle"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Molecular chaperones of the Hsp70 family assist in the assembly of 20S proteasomes
    (Elsevier, 2017-04) Hammack, Lindsay J.; Firestone, Kyle; Chang, William; Kusmierczyk, Andrew R.; Department of Biology, School of Science
    The eukaryotic 26S proteasome is a large protease comprised of two major sub assemblies, the 20S proteasome, or core particle (CP), and the 19S regulatory particle (RP). Assembly of the CP and RP is assisted by an expanding list of dedicated assembly factors. For the CP, this includes Ump1 and the heterodimeric Pba1–Pba2 and Pba3–Pba4 proteins. It is not known how many additional proteins that assist in proteasome biogenesis remain to be discovered. Here, we demonstrate that two members of the Hsp70 family in yeast, Ssa1 and Ssa2, play a direct role in CP assembly. Ssa1 and Ssa2 interact genetically and physically with proteasomal components. Specifically, they associate tightly with known CP assembly intermediates, but not with fully assembled CP, through an extensive purification protocol. And, in yeast lacking both Ssa1 and Ssa2, specific defects in CP assembly are observed.
  • Thumbnail Image
    Item
    YPL260W, a high-copy suppressor of a copper-sensitive phenotype in yeast, is linked to DNA repair and proteasome function
    (Elsevier, 2015-11-27) Firestone, Kyle; Awonusi, Damilola; Panfair, Dilrajkaur; Roland, Derrick; Ramamurthy, Aishwarya; Kusmierczyk, Andrew R.; Department of Biology, School of Science
    The ubiquitin–proteasome system directly impacts the metabolism of heavy metals and yeast has become an important model in understanding this interplay. We demonstrate that yeast mutants with defects in proteasome function are able to tolerate elevated levels of copper. In the course of our analysis, we isolate a yeast mutant that not only negates this copper tolerance in proteasome mutants, but renders yeast exquisitely sensitive to this metal. To better understand the nature of the defect, we carry out a plasmid-based genetic screen to identify high-copy suppressors of this strong copper sensitivity. We identify four genes not previously known to be associated with copper metabolism: CDC53, PSP1, YNL200C, and YPL260W. The latter is a highly conserved fungal gene of no known function. Here, we undertake the first characterization of YPL260W. We demonstrate YPL260W to have a role in bleomycin tolerance with links to DNA repair and proteasome function.