Mammalian Prenylcysteine Carboxyl Methyltransferase Is in the Endoplasmic Reticulum*
- Qun Dai‡,
- Edwin Choy‡,
- Vi Chiu‡,
- Julia Romano§,
- Sandra R. Slivka¶‖,
- Susan A. Steitz¶**,
- Susan Michaelis§ and
- Mark R. Philips‡‡
- From the ‡Departments of Medicine and Cell Biology, New York University School of Medicine, New York, New York 10016,§Department of Cell Biology and Anatomy, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, and¶Tanabe Research Laboratories, USA, Inc., San Diego, California 92121
Abstract
Prenylcysteine carboxyl methyltransferase (pcCMT) is the third of three enzymes that posttranslationally modify C-terminal CAAX motifs and thereby target CAAXproteins to the plasma membrane. Here we report the molecular characterization and subcellular localization of the first mammalian (human myeloid) pcCMT. The deduced amino acid sequence of mammalian pcCMT predicts a multiple membrane-spanning protein with homologies to the yeast pcCMT, STE14, and the mammalian band 3 anion transporter. The human gene complemented a ste14 mutant. pcCMT mRNAs were ubiquitously expressed in human tissues. An anti-pcCMT antiserum detected a 33-kDa protein in myeloid cell membranes. Ectopically expressed recombinant pcCMT had enzymatic activity identical to that observed in neutrophil membranes. Mammalian pcCMT was not expressed at the plasma membrane but rather restricted to the endoplasmic reticulum. Thus, the final enzyme in the sequence that modifies CAAX motifs is located in membranes topologically removed from the CAAX protein target membrane.
Footnotes
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↵* This work was supported by grants from the National Institutes of Health (AI36224 and GM55279 (to M. R. P.) and GM41223 (to S. M.)) and from the American Heart Association and the Council for Tobacco Research (to M. R. P).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.
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↵‖ Present address: Trega, 3550 General Atomics Ct., San Diego, CA 92121.
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↵** Present address: Dept. of Pathology, University of Washington, 1959 NE Pacific St., Seattle, WA.
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↵‡ To whom correspondence should be addressed. Tel.: 212-263-6403; Fax: 212-263-8804; E-mail: mark.philips{at}mcccm.med.nyu.edu.
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↵1 The abbreviations used are: CAAX, tetrapeptide with sequence cysteine-(usually aliphatic amino acid)2-any amino acid; pcCMT, prenylcysteine carboxyl methyltransferase; GFP, green fluorescent protein; ER, endoplasmic reticulum; EST, expressed sequence tag; ORF, open reading frame; PCR, polymerase chain reaction; AFC,N-acetyl-S-trans,trans-farnesyl-l-cysteine; GDI, guanine nucleotide dissociation inhibitor; bp, base pair; kb, kilobase; CHO, Chinese hamster ovary; GTPγS, guanosine 5′-3-O-(thio)triphosphate; AGGC,N-acetyl-S-all-trans-geranylgeranyl-l-cysteine; Me2SO, dimethyl sulfoxide.
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↵2 J. Romano, W. Schmidt, and S. Michaelis, submitted for publication.
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↵3 M. Philips and T. Oei, unpublished results.
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↵4 J. Romano, W. Schmidt, and S. Michaelis, submitted for publication.
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- Received February 25, 1998.
- Revision received April 8, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
