Isoprenylcysteine Carboxyl Methyltransferase Deficiency in Mice

doi:10.1074/jbc.C000831200

Isoprenylcysteine Carboxyl Methyltransferase Deficiency in Mice*

From the ^‡Gladstone Institute of Cardiovascular Disease, the ^§Cardiovascular Research Institute, and the ^‖Department of Medicine, University of California, San Francisco, California 94141-9100; the ^**Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27710-3686; and the ^‡Molecular Genetics Section, Division of Biomedical Sciences, Imperial College School of Medicine, Sir Alexander Fleming Building, London, SW7 2AZ, United Kingdom

Abstract

After isoprenylation, Ras and other CAAX proteins undergo endoproteolytic processing by Rce1 and methylation of the isoprenylcysteine by Icmt (isoprenylcysteine carboxyl methyltransferase). We reported previously thatRce1-deficient mice died during late gestation or soon after birth. We hypothesized that Icmt deficiency might cause a milder phenotype, in part because of reports suggesting the existence of more than one activity for methylating isoprenylated proteins. To address this hypothesis and also to address the issue of other methyltransferase activities, we generatedIcmt-deficient mice. Contrary to our expectation,Icmt deficiency caused a more severe phenotype thanRce1 deficiency, with virtually all of the knockout embryos (Icmt−/−) dying by mid-gestation. An analysis of chimeric mice produced from Icmt−/− embryonic stem cells showed that the Icmt−/− cells retained the capacity to contribute to some tissues (e.g. skeletal muscle) but not to others (e.g. brain). Lysates from Icmt−/− embryos lacked the ability to methylate either recombinant K-Ras or small molecule substrates (e.g. N-acetyl-S-geranylgeranyl-l-cysteine). In addition, Icmt−/− cells lacked the ability to methylate Rab proteins. Thus, Icmt appears to be the only enzyme participating in the carboxyl methylation of isoprenylated proteins.

Footnotes

↵* This work was supported in part by National Institutes of Health Grants HL41633 and AG15451 (to S. G. Y.) and GM46372 (to P. J. C), a Wellcome Trust Programme Grant (to M. C. S.), and grant awards from the University of California Tobacco-related Disease Research Program (to M. O. B. and S. G. Y.).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.
↵¶ To whom correspondence should be addressed: Gladstone Institute of Cardiovascular Disease, P. O. Box 419100, San Francisco, CA 94141-9100. Tel.: 415-695-3774; Fax: 415-285-5632; E-mail: mbergo@gladstone.ucsf.edu.
↵§§ Recipient of a Ph.D. Student Award from Fundação Ciência e Tecnologia of Portugal.
Published, JBC Papers in Press, December 19, 2000, DOI 10.1074/jbc.C000831200
↵2 Although no Southern blots were performed on skin DNA, the percentage of brown fur on the chimeric animals suggested that Icmt-deficient ES cells are fairly robust in contributing to the formation of the hair follicles.
↵3 Zmpste24 is the mouse ortholog ofAFC1 (STE24) in Saccharomyces cerevisiae (2, 27, 28).
↵4 G. Leung, M. Bergo, and S. Young, unpublished observations.
Abbreviations:

CAAX

C-terminal motif consisting of Cys followed by two aliphatic residues (A) and any amino acid (X)

Icmt

isoprenylcysteine carboxyl methyltransferase

ES

embryonic stem

AGGC

N-acetyl-S-geranylgeranyl-l-cysteine

AFC

N-acetyl-S-farnesyl-l-cysteine

GGPP

geranylgeranylpyrophosphate

REP

Rab escort protein

RabGGTase

Rab geranylgeranyltransferase

kb

kilobases
- Received November 26, 2000.
- Revision received December 14, 2000.
The American Society for Biochemistry and Molecular Biology, Inc.