Characterization of an ADP-ribosylation Factor-like 1 Protein inSaccharomyces cerevisiae*
- Fang-Jen S. Lee‡§,
- Chun-Fang Huang‡,
- Wei-Luen Yu‡,
- Leh-Miauh Buu‡,
- Ching-Yi Lin‡,
- Min-Chuan Huang‡,
- Joel Moss¶ and
- Martha Vaughan¶
- From the ‡Institute of Molecular Medicine, School of Medicine, National Taiwan University, Taipei, Taiwan, Republic of China and the ¶Pulmonary-Critical Care Medicine Branch, NHLBI, National Institutes of Health, Bethesda, Maryland 20892-1434
Abstract
ADP-ribosylation factors (ARFs) are highly conserved ∼20-kDa guanine nucleotide-binding proteins that enhance the ADP-ribosyltransferase activity of cholera toxin and are believed to participate in vesicular transport in both exocytic and endocytic pathways. Several ARF-like proteins (ARLs) have been cloned fromDrosophila, rat, and human; however, the biological functions of ARLs are unknown. We have identified a yeast gene (ARL1) encoding a protein that is structurally related (>60% identical) to human, rat, and Drosophila ARL1. Biochemical analyses of purified recombinant yeast ARL1 (yARL1) protein revealed properties similar to those ARF and ARL1 proteins, including the ability to bind and hydrolyze GTP. Like other ARLs, recombinant yARL1 protein did not stimulate cholera toxin-catalyzed auto-ADP-ribosylation. yARL1 was not recognized by antibodies against mammalian ARLs or yeast ARFs. Anti-yARL1 antibodies did not cross-react with yeast ARFs, but did react with human ARLs. On subcellular fractionation, yARL1, similar to yARF1, was localized to the soluble fraction. The amino terminus of yARL1, like that of ARF, was myristoylated. Unlike Drosophila Arl1, yeastARL1 was not essential for cell viability. Like rat ARL1, yARL1 might be associated in part with the Golgi complex. However, yARL1 was not required for endoplasmic reticulum-to-Golgi protein transport, and it may offer an opportunity to define an ARL function in another kind of vesicular trafficking, such as the regulated secretory pathway.
Footnotes
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↵* This work was supported by Grant NSC-86-2314-B-002-203 from the National Science Council, Republic of China (to F.-J. S. L.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The nucleotide sequence reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number U89332. The complete sequence of this fragment had been deposited in data bases by the Yeast Genome Project (accession number z36033, chromosome II, ORF:YBR164c).
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↵§ To whom correspondence should be addressed: Inst. of Molecular Medicine, School of Medicine, National Taiwan University, 7 Chung Shan South Rd., Taipei, Taiwan. Tel.: 886-2-397-0800 (ext. 5730); Fax: 886-2-321-0977; E-mail: fangjen{at}ha.mc.ntu.edu.tw.
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↵1 The abbreviations used are: ARFs, ADP-ribosylation factors; yARF, recombinant yeast ARF (S. cerevisiae); ARLs, ARF-like proteins; hARL, human ARL; rARL, rat ARL; ER, endoplasmic reticulum; PCR, polymerase chain reaction; kb, kilobase pair(s); PAGE, polyacrylamide gel electrophoresis; HA, hemagglutinin; GTPγS, guanosine 5′-O-(3thiotriphosphate).
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↵2 F.-J. S. Lee, C.-F. Huang, W.-L. Yu, L.-M. Buu, C.-Y. Lin, J. Moss, and M. Vaughan, unpublished data.
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- Received April 16, 1997.
- Revision received August 4, 1997.
