A role for N-myristoylation in protein targeting: NADH-cytochrome b5 reductase requires myristic acid for association with outer mitochondrial but not ER membranes

doi:10.1083/jcb.135.6.1501

This Article

	Full Text (PDF, 3604K)
	Alert me when this article is cited
	Citation Map

Services

	Email this article
	Similar articles in this journal
	Similar articles in PubMed
	Alert me to new content in the JCB
	Download to citation manager

Citing Articles

	Citing Articles via HighWire
	Citing Articles via CrossRef
	Citing Articles via Google Scholar

Google Scholar

	Articles by Borgese, N.
	Articles by Bassetti, M.
	Search for Related Content

PubMed

	PubMed Citation
	Articles by Borgese, N.
	Articles by Bassetti, M.


Social Bookmarking

	What's this?

ARTICLES

A role for N-myristoylation in protein targeting: NADH-cytochrome b5 reductase requires myristic acid for association with outer mitochondrial but not ER membranes

N Borgese, D Aggujaro, P Carrera, G Pietrini and M Bassetti
Consiglio Nazionale delle Ricerche Cellular and Molecular Pharmacology Center, Department of Pharmacology, University of Milan, Italy. Nica@Farmal.csfic.mi.cnr.it

N-myristoylation is a cotranslational modification involved in protein- protein interactions as well as in anchoring polypeptides to phospholipid bilayers; however, its role in targeting proteins to specific subcellular compartments has not been clearly defined. The mammalian myristoylated flavoenzyme NADH-cytochrome b5 reductase is integrated into ER and mitochondrial outer membranes via an anchor containing a stretch of 14 uncharged amino acids downstream to the NH2- terminal myristoylate glycine. Since previous studies suggested that the anchoring function could be adequately carried out by the 14 uncharged residues, we investigated a possible role for myristic acid in reductase targeting. The wild type (wt) and a nonmyristoylatable reductase mutant (gly2-->ala) were stably expressed in MDCK cells, and their localization was investigated by immunofluorescence, immuno-EM, and cell fractionation. By all three techniques, the wt protein localized to ER and mitochondria, while the nonmyristoylated mutant was found only on ER membranes. Pulse-chase experiments indicated that this altered steady state distribution was due to the mutant's inability to target to mitochondria, and not to its enhanced instability in that location. Both wt and mutant reductase were resistant to Na2CO3 extraction and partitioned into the detergent phase after treatment of a membrane fraction with Triton X-114, demonstrating that myristic acid is not required for tight anchoring of reductase to membranes. Our results indicate that myristoylated reductase localizes to ER and mitochondria by different mechanisms, and reveal a novel role for myristic acid in protein targeting.
Add to CiteULike CiteULike Add to Complore Complore Add to Connotea Connotea Add to Del.icio.us Del.icio.us Add to Digg Digg Facebook Add to Reddit Reddit Add to Technorati Technorati Twitter What's this?

This article has been cited by other articles:

Lu, B., Poirier, C., Gaspar, T., Gratzke, C., Harrison, W., Busija, D., Matzuk, M. M., Andersson, K.-E., Overbeek, P. A., Bishop, C. E. (2008). A Mutation in the Inner Mitochondrial Membrane Peptidase 2-Like Gene (Immp2l) Affects Mitochondrial Function and Impairs Fertility in Mice. Biol. Reprod. 78: 601-610 [Abstract] [Full Text]
Sprocati, T., Ronchi, P., Raimondi, A., Francolini, M., Borgese, N. (2006). Dynamic and reversible restructuring of the ER induced by PDMP in cultured cells. J. Cell Sci. 119: 3249-3260 [Abstract] [Full Text]
Raffaello, A., Laveder, P., Romualdi, C., Bean, C., Toniolo, L., Germinario, E., Megighian, A., Danieli-Betto, D., Reggiani, C., Lanfranchi, G. (2006). Denervation in murine fast-twitch muscle: short-term physiological changes and temporal expression profiling. Physiol. Genomics 25: 60-74 [Abstract] [Full Text]
Colombo, S., Longhi, R., Alcaro, S., Ortuso, F., Sprocati, T., Flora, A., Borgese, N. (2005). N-myristoylation determines dual targeting of mammalian NADH-cytochrome b(5) reductase to ER and mitochondrial outer membranes by a mechanism of kinetic partitioning. JCB 168: 735-745 [Abstract] [Full Text]
Zhu, H., Larade, K., Jackson, T. A., Xie, J., Ladoux, A., Acker, H., Berchner-Pfannschmidt, U., Fandrey, J., Cross, A. R., Lukat-Rodgers, G. S., Rodgers, K. R., Bunn, H. F. (2004). NCB5OR Is a Novel Soluble NAD(P)H Reductase Localized in the Endoplasmic Reticulum. J. Biol. Chem. 279: 30316-30325 [Abstract] [Full Text]
Bulbarelli, A., Sprocati, T., Barberi, M., Pedrazzini, E., Borgese, N. (2002). Trafficking of tail-anchored proteins: transport from the endoplasmic reticulum to the plasma membrane and sorting between surface domains in polarised epithelial cells. J. Cell Sci. 115: 1689-1702 [Abstract] [Full Text]
Le Blanc, I., Blot, V., Bouchaert, I., Salamero, J., Goud, B., Rosenberg, A. R., Dokhelar, M.-C. (2002). Intracellular Distribution of Human T-Cell Leukemia Virus Type 1 Gag Proteins Is Independent of Interaction with Intracellular Membranes. J. Virol. 76: 905-911 [Abstract] [Full Text]
Borgese, N., Gazzoni, I., Barberi, M., Colombo, S., Pedrazzini, E. (2001). Targeting of a Tail-anchored Protein to Endoplasmic Reticulum and Mitochondrial Outer Membrane by Independent but Competing Pathways. Mol. Biol. Cell 12: 2482-2496 [Abstract] [Full Text]
Zha, J., Weiler, S., Oh, K. J., Wei, M. C., Korsmeyer, S. J. (2000). Posttranslational N-Myristoylation of BID as a Molecular Switch for Targeting Mitochondria and Apoptosis. Science 290: 1761-1765 [Abstract] [Full Text]
Pedrazzini, E., Villa, A., Longhi, R., Bulbarelli, A., Borgese, N. (2000). Mechanism of Residence of Cytochrome B(5), a Tail-Anchored Protein, in the Endoplasmic Reticulum. JCB 148: 899-914 [Abstract] [Full Text]
Sarkar, S. N., Bandyopadhyay, S., Ghosh, A., Sen, G. C. (1999). Enzymatic Characteristics of Recombinant Medium Isozyme of 2'-5' Oligoadenylate Synthetase. J. Biol. Chem. 274: 1848-1855 [Abstract] [Full Text]
Fukuchi-Mizutani, M., Mizutani, M., Tanaka, Y., Kusumi, T., Ohta, D. (1999). Microsomal Electron Transfer in Higher Plants: Cloning and Heterologous Expression of NADH-Cytochrome b5 Reductase from Arabidopsis. Plant Physiol. 119: 353-362 [Abstract] [Full Text]
Bulbarelli, A., Valentini, A., DeSilvestris, M., Cappellini, M. D., Borgese, N. (1998). An Erythroid-Specific Transcript Generates the Soluble Form of NADH-Cytochrome b5 Reductase in Humans. Blood 92: 310-319 [Abstract] [Full Text]
Belcourt, M. F., Hodnick, W. F., Rockwell, S., Sartorelli, A. C. (1998). The Intracellular Location of NADH:Cytochrome b5 Reductase Modulates the Cytotoxicity of the Mitomycins to Chinese Hamster Ovary Cells. J. Biol. Chem. 273: 8875-8881 [Abstract] [Full Text]
Matsuzawa, A., Hattori, K., Aoki, J., Arai, H., Inoue, K. (1997). Protection against Oxidative Stress-induced Cell Death by Intracellular Platelet-activating Factor-Acetylhydrolase II. J. Biol. Chem. 272: 32315-32320 [Abstract] [Full Text]
Glover, C. J., Hartman, K. D., Felsted, R. L. (1997). Human N-Myristoyltransferase Amino-terminal Domain Involved in Targeting the Enzyme to the Ribosomal Subcellular Fraction. J. Biol. Chem. 272: 28680-28689 [Abstract] [Full Text]