Rad4 regulates protein turnover at a postubiquitylation step.
The ubiquitin (Ub)-binding protein Rad23 plays an important role in facilitating the transfer of substrates to the proteasome. However, the mechanism underlying Rad23's function in proteolysis remains unknown. Here, we demonstrate that Rad4, a Rad23-binding protein, also regulates ubiquitylated substrate turnover. Rad4 was known previously only as a key repair factor that directly recognizes DNA damage and initiates DNA repair. Our results, however, reveal a novel function of Rad4. We found that Rad4 and Rad23 share several common substrates. Substrates in rad4Delta cells are ubiquitylated, indicating that Rad4 regulates a postubiquitylation event. Moreover, we found that Rad4 participates in the Rad23-Ufd2 pathway, but not the Rad23-Png1 pathway, consistent with previous findings that Png1 and Rad4 or Ufd2 form separate Rad23 complexes. The Rad4-binding domain is crucial for the functioning of Rad23 in degradation, suggesting that Rad4 and Rad23 work together in proteolysis. It is interesting to note that upon DNA damage, Rad4 becomes concentrated in the nucleus and degradation of the nonnuclear protein Pex29 is compromised, further suggesting that Rad4 may influence the coordination of various cellular processes. Our findings will help to unravel the detailed mechanisms underlying the roles of Rad23 and Rad4 in proteolysis and also the interplay between DNA repair and proteolysis.
Li
Yue
Y
Department of Molecular Medicine, Institute of Biotechnology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78245, USA.
Yan
Jing
J
Kim
Ikjin
I
Liu
Chang
C
Huo
Keke
K
Rao
Hai
H
D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D019913:Nuclear Localization Signals / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D017384:Sequence Deletion; D013379:Substrate Specificity; D025801:Ubiquitin / Q000378:metabolism; D054875:Ubiquitination
D004268:DNA-Binding Proteins; D006023:Glycoproteins; D019913:Nuclear Localization Signals; C493797:Rad4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D046988:Proteasome Endopeptidase Complex