Browsing by Author "London, MK"
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- Regulatory mechanisms controlling biogenesis of ubiquitin and the proteasomePublication . London, MK; Keck, BI; Ramos, PC; Dohmen, RHAnalysis of several Saccharomyces cerevisiae ump mutants with defects in ubiquitin (Ub)-mediated proteolysis yielded insights into the regulation of the polyubiquitin gene UB14 and of proteasome genes. High-molecular weight Ub-protein conjugates accumulated in ump mutants with impaired proteasome function with a concomitant decrease in the amount of free Ub. In these mutants, transcriptional induction of UB14 was depending in part on the transcription factor Rpn4. Deletion of UB14 partially suppressed the growth defects of ump1 mutants, indicating that accumulation of polyubiquitylated proteins is deleterious to cell growth. Transcription of proteasome subunit genes was induced in ump mutants affecting the proteasome, as well as under conditions that mediate DNA damage or the formation of abnormal proteins. This induction required the transcriptional activator Rpn4. Elevated Rpn4 levels in proteasome-deficient mutants or as a response to abnormal proteins were due to increased metabolic stability. Up-regulation of proteasome genes in response to DNA damage, in contrast, is shown to operate via induction of RPN4 transcription. (C) 2004 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
- Role of C-terminal extensions of subunits beta 2 and beta 7 in assembly and activity of eukaryotic proteasomesPublication . Ramos, Paula C; Marques, AJ; London, MK; Dohmen, RJA close inspection of the crystal structure of the yeast 20 S proteasome revealed that a prominent connection between the two beta-rings is mediated by the subunit beta7/ Pre4. Its C-terminal extension intercalates between the beta1/Pre3 and beta2/Pup1 subunits on the opposite ring. We show that the interactions promoted by the beta7/Pre4 tail are important to facilitate the formation of 20 S particles from two half-proteasome precursor complexes and/or to stabilize mature 20 S proteasomes. The deletion of 19 residues from the beta7/Pre4 C terminus leads to an accumulation of half-proteasome precursor complexes containing the maturation factor Ump1. The C-terminal extension of beta7/Pre4, which forms several hydrogen bonds with beta1/Pre3, is in addition required for the post-acidic activity mediated by the latter subunit. Deletion of the C-terminal tail of beta7/Pre4 results in an inhibition of beta1/Pre3 propeptide processing and abrogation of post-acidic activity. Our data obtained with yeast strains that expressed the mature form of Pre3 lacking its propeptide suggest that interactions between the Pre4 C terminus and Pre3 stabilize a conformation of its active site, which is essential for post-acidic activity. Deletion of the C-terminal extension of beta2/Pup1, which wraps around beta3/Pup3 within the same beta-ring, is lethal, indicating that this extension serves an essential function in proteasome assembly or stability.