Bacterial and animal cells selectively degrade proteins with abnormal conformations. This process helps prevent the intracellular accumulation of unfolded proteins in genetic diseases, during heat shock, and with aging. Molecular chaperones, in addition to catalyzing protein folding and translocation, play essential roles in this degradative process. During the current funding period the PI has shown that in E. coli the chaperones DnaK, DnaJ, and GrpE are necessary for the rapid degradation of certain unfolded model polypeptides while GroEL, GroES, and Trigger Factor function together in the breakdown of other (abnormal) polypeptides. These chaperones associate with the substrates and appear to release or present them in a conformation that facilitates proteolytic digestion. In yeast the PI has shown that the DnaJ homolog Ydj1 and Hsp70 of the SSA family are essential for ubiquitin conjugation to abnormal proteins and certain regulatory proteins by binding and enhancing the susceptibility for ubiquitination. The current grant is comprised of four specific aims.
Specific Aim A will continue studies on the roles of GroEL/ES/TF in the degradation of the recombinant model substrate CRAG by delineating discrete steps in the initial fragmentation. These studies will combine proteolytic susceptibility studies, mass spec analysis of peptide products, and direct structure determination by EM tomography in collaboration Wolfgang Baumeister. Other studies will test whether the model CRAG substrate is degraded through an analogous chaperone dependent pathway in yeast, allowing the use of yeast genetics to dissect steps.
Specific Aim B will focus on clarifying the functions of DnaK and DnaJ in the degradation of the unfolded protein PhoA in E. coli. These studies will use His tagged PhoA and protease dificient strains to isolate intact substrate-chaperone complexes.
Specific Aim C will determine the roles of the DnaJ and Hsp70 families in the ubiquitin-proteasome pathway in yeast using model substrates carrying defined degradation signals. This line of investigation will be extended to defined cell free studies as well as studies of the potential role of Sis1 in binding to ubiquitin conjugates prior to degradation by the proteasome.
Specific Aim D will define the chaperone like function of the PAN and HslU ATPases which support protein breakdown by proteasome homologs in archeabacteria and E. coli.
|Sha, Zhe; Schnell, Helena M; Ruoff, Kerstin et al. (2018) Rapid induction of p62 and GABARAPL1 upon proteasome inhibition promotes survival before autophagy activation. J Cell Biol 217:1757-1776|
|Kim, Hyoung Tae; Goldberg, Alfred L (2018) UBL domain of Usp14 and other proteins stimulates proteasome activities and protein degradation in cells. Proc Natl Acad Sci U S A 115:E11642-E11650|
|VerPlank, Jordan J S; Lokireddy, Sudarsanareddy; Feltri, M Laura et al. (2018) Impairment of protein degradation and proteasome function in hereditary neuropathies. Glia 66:379-395|
|Kim, Hyoung Tae; Goldberg, Alfred L (2017) The deubiquitinating enzyme Usp14 allosterically inhibits multiple proteasomal activities and ubiquitin-independent proteolysis. J Biol Chem 292:9830-9839|
|Weyburne, Emily S; Wilkins, Owen M; Sha, Zhe et al. (2017) Inhibition of the Proteasome ?2 Site Sensitizes Triple-Negative Breast Cancer Cells to ?5 Inhibitors and Suppresses Nrf1 Activation. Cell Chem Biol 24:218-230|
|VerPlank, Jordan J S; Goldberg, Alfred L (2017) Regulating protein breakdown through proteasome phosphorylation. Biochem J 474:3355-3371|
|Edison, Natalia; Curtz, Yael; Paland, Nicole et al. (2017) Degradation of Bcl-2 by XIAP and ARTS Promotes Apoptosis. Cell Rep 21:442-454|
|Volodin, Alexandra; Kosti, Idit; Goldberg, Alfred Lewis et al. (2017) Myofibril breakdown during atrophy is a delayed response requiring the transcription factor PAX4 and desmin depolymerization. Proc Natl Acad Sci U S A 114:E1375-E1384|
|Kuo, Chueh-Ling; Goldberg, Alfred Lewis (2017) Ubiquitinated proteins promote the association of proteasomes with the deubiquitinating enzyme Usp14 and the ubiquitin ligase Ube3c. Proc Natl Acad Sci U S A 114:E3404-E3413|
|Collins, Galen Andrew; Goldberg, Alfred L (2017) The Logic of the 26S Proteasome. Cell 169:792-806|
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