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Ubiquitin conjugation by the N-end rule pathway and mRNAs for its components increase in muscles of diabetic rats

Lecker, Stewart H. and Solomon, Vered and Price, S. Russ and Kwon, Yong Tae and Mitch, William E. and Goldberg, Alfred L. (1999) Ubiquitin conjugation by the N-end rule pathway and mRNAs for its components increase in muscles of diabetic rats. Journal of Clinical Investigation, 104 (10). pp. 1411-1420. ISSN 0021-9738. http://resolver.caltech.edu/CaltechAUTHORS:20111220-100811409

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Abstract

Insulin deficiency (e.g., in acute diabetes or fasting) is associated with enhanced protein breakdown in skeletal muscle leading to muscle wasting. Because recent studies have suggested that this increased proteolysis is due to activation of the ubiquitin-proteasome (Ub-proteasome) pathway, we investigated whether diabetes is associated with an increased rate of Ub conjugation to muscle protein. Muscle extracts from streptozotocin-induced insulin-deficient rats contained greater amounts of Ub-conjugated proteins than extracts from control animals and also 40–50% greater rates of conjugation of ^(125)I-Ub to endogenous muscle proteins. This enhanced Ub-conjugation occurred mainly through the N-end rule pathway that involves E2_(14k) and E3α. A specific substrate of this pathway, α-lactalbumin, was ubiquitinated faster in the diabetic extracts, and a dominant negative form of E2_(14k) inhibited this increase in ubiquitination rates. Both E2_(14k) and E3α were shown to be rate-limiting for Ub conjugation because adding small amounts of either to extracts stimulated Ub conjugation. Furthermore, mRNA for E2_(14k) and E3α (but not E1) were elevated 2-fold in muscles from diabetic rats, although no significant increase in E2_(14k) and E3α content could be detected by immunoblot or activity assays. The simplest interpretation of these results is that small increases in both E2_(14k) and E3α in muscles of insulin-deficient animals together accelerate Ub conjugation and protein degradation by the N-end rule pathway, the same pathway activated in cancer cachexia, sepsis, and hyperthyroidism.


Item Type:Article
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URLURL TypeDescription
http://dx.doi.org/10.1172/JCI7300 DOIUNSPECIFIED
http://www.jci.org/articles/view/7300PublisherUNSPECIFIED
Additional Information:© 1999 The American Society for Clinical Investigation. Received for publication May 10, 1999, and accepted in revised form October 6, 1999. This work is supported in part by grants from the National Space Biomedical Research Institute (to A.L. Goldberg), National Institutes of Health (GM-46147 to A.L. Goldberg, DK-37175 and HL-45317 to W.E. Mitch, and DK-50740 to S.R. Price), and the Muscular Dystrophy Foundation (to A.L. Goldberg). S.H. Lecker is a Physician Postdoctoral Fellow of the Howard Hughes Medical Institute. We are grateful to Jackie Pierce, Margaret Read, and Vincent Chau for providing the dominant negative E2s, Chikara Miyamoto for providing the E1 overproducer, Art Haas for providing the antibody to E214k, and Aaron Ciechanover for providing the antiserum to E1. We also thank O. Kandror and E. Tarcsa for their helpful discussions and critical reading of the manuscript.
Funders:
Funding AgencyGrant Number
National Space Biomedical Research Institute UNSPECIFIED
NIHGM-46147
NIHDK-37175
NIHHL-45317
NIHDK-50740
Muscular Dystrophy Foundation UNSPECIFIED
Record Number:CaltechAUTHORS:20111220-100811409
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20111220-100811409
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:28530
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:20 Dec 2011 18:38
Last Modified:26 Dec 2012 14:38

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