Kwon, Yong Tae and Kashina, Anna S. and Varshavsky, Alexander (1999) Alternative Splicing Results in Differential Expression, Activity, and Localization of the Two Forms of Arginyl-tRNA-Protein Transferase, a Component of the N-End Rule Pathway. Molecular and Cellular Biology, 19 (1). pp. 182-193. ISSN 0270-7306. PMCID PMC83877. doi:10.1128/mcb.19.1.182. https://resolver.caltech.edu/CaltechAUTHORS:KWOmcb99
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Abstract
The N-end rule relates the in vivo half-life of a protein to the identity of its N-terminal residue. The underlying ubiquitin-dependent proteolytic system, called the N-end rule pathway, is organized hierarchically: N-terminal aspartate and glutamate (and also cysteine in metazoans) are secondary destabilizing residues, in that they function through their conjugation, by arginyl-tRNA-protein transferase (R-transferase), to arginine, a primary destabilizing residue. We isolated cDNA encoding the 516-residue mouse R-transferase, ATE1p, and found two species, termed Ate1-1 and Ate1-2. The Ate1 mRNAs are produced through a most unusual alternative splicing that retains one or the other of the two homologous 129-bp exons, which are adjacent in the mouse Ate1 gene. Human ATE1 also contains the alternative 129-bp exons, whereas the plant (Arabidopsis thaliana) and fly (Drosophila melanogaster) Ate1 genes encode a single form of ATE1p. A fusion of ATE1-1p with green fluorescent protein (GFP) is present in both the nucleus and the cytosol, whereas ATE1-2p-GFP is exclusively cytosolic. Mouse ATE1-1p and ATE1-2p were examined by expressing them in ate1Delta Saccharomyces cerevisiae in the presence of test substrates that included Asp-beta gal (beta -galactosidase) and Cys-beta gal. Both forms of the mouse R-transferase conferred instability on Asp-beta gal (but not on Cys-beta gal) through the arginylation of its N-terminal Asp, the ATE1-1p enzyme being more active than ATE1-2p. The ratio of Ate1-1 to Ate1-2 mRNA varies greatly among the mouse tissues; it is ~0.1 in the skeletal muscle, ~0.25 in the spleen, ~3.3 in the liver and brain, and ~10 in the testis, suggesting that the two R-transferases are functionally distinct.
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Additional Information: | © 1999, American Society for Microbiology. Received 13 August 1998/Returned for modification 21 September 1998/Accepted 6 October 1998 The first two authors contributed equally to this work. We thank Gary Hathaway of the Caltech Microchemistry Facility for the sequencing of X-beta gal proteins. We are grateful to Hai Rao, Glenn Turner, Fangyong Du, and Lawrence Peck for helpful suggestions and to Fangyong Du, Federico Navarro-Garcia, Hai Rao, and Youming Xie for comments on the manuscript. This work was supported by grants DK39520 and GM31530 to A.V. from the National Institutes of Health. | |||||||||
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Issue or Number: | 1 | |||||||||
PubMed Central ID: | PMC83877 | |||||||||
DOI: | 10.1128/mcb.19.1.182 | |||||||||
Record Number: | CaltechAUTHORS:KWOmcb99 | |||||||||
Persistent URL: | https://resolver.caltech.edu/CaltechAUTHORS:KWOmcb99 | |||||||||
Usage Policy: | No commercial reproduction, distribution, display or performance rights in this work are provided. | |||||||||
ID Code: | 6789 | |||||||||
Collection: | CaltechAUTHORS | |||||||||
Deposited By: | Archive Administrator | |||||||||
Deposited On: | 21 Dec 2006 | |||||||||
Last Modified: | 08 Nov 2021 20:37 |
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