HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 280, Issue 14, 14097-14104, April 8, 2005

This Article Full Text Full Text (PDF) All Versions of this Article: 280/14/14097    most recent M500147200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Loewen, C. J. R. Articles by Levine, T. P. Search for Related Content PubMed PubMed Citation Articles by Loewen, C. J. R. Articles by Levine, T. P. Social Bookmarking                      What's this?

A Highly Conserved Binding Site in Vesicle-associated Membrane Protein-associated Protein (VAP) for the FFAT Motif of Lipid-binding Proteins^*

Christopher J. R. Loewen and Timothy P. Levine

From the Division of Cell Biology, Institute of Ophthalmology, University College London, Bath Street, London EC1V 9EL, United Kingdom

A variety of lipid-binding proteins contain a recently described motif, designated FFAT (two phenylalanines in an acidic tract), which binds to vesicle-associated-membrane protein-associated protein (VAP). VAP is a conserved integral membrane protein of the endoplasmic reticulum that contains at its amino terminus a domain related to the major sperm protein of nematode worms. Here we have studied the FFAT-VAP interaction in Saccharomyces cerevisiae, where the VAP homologue Scs2 regulates phospholipid metabolism via an interaction with the FFAT motif of Opi1. By introducing mutations at random into Scs2, we found that mutations that abrogated binding to FFAT were clustered in the most highly conserved region. Using site-directed mutagenesis, we identified several critical residues, including two lysines widely separated in the primary sequence. By examining all other conserved basic residues, we identified a third residue that was moderately important for binding FFAT. Modeling VAP on the known structure of major sperm protein showed that the critical residues form a patch on a positively charged face of the protein. In vivo functional studies of SCS22, a second SCS2-like gene in S. cerevisiae, showed that SCS2 was the dominant gene in the regulation of Opi1, with a minor contribution from SCS22. We then established that reduction in the affinity of Scs2 mutants for FFAT correlated well with loss of function, indicating the importance of these residues for binding FFAT motifs. Finally, we found that human VAP-A could substitute for Scs2 but that it functioned poorly, suggesting that other factors modulate the binding of Scs2 to proteins with FFAT motifs.

Received for publication, January 5, 2005 , and in revised form, January 21, 2005.

^* This work was supported by Grant 31/C15982 from the Biotechnology and Biological Sciences Research Council, Wellcome Trust Grant 060537, the Canadian Institutes of Health Research (to C. J. R. L.), and Fight For Sight. The costs of publication of this article were defrayed in part by the payment of page charges. This 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. Tel.: 44-20-7608-4027; Fax: 44-20-7608-4034; E-mail: tim.levine{at}ucl.ac.uk.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				D. C. Prosser, D. Tran, P.-Y. Gougeon, C. Verly, and J. K. Ngsee FFAT rescues VAPA-mediated inhibition of ER-to-Golgi transport and VAPB-mediated ER aggregation J. Cell Sci., September 15, 2008; 121(18): 3052 - 3061. [Abstract] [Full Text] [PDF]

				G.-S. Han, L. O'Hara, S. Siniossoglou, and G. M. Carman Characterization of the Yeast DGK1-encoded CTP-dependent Diacylglycerol Kinase J. Biol. Chem., July 18, 2008; 283(29): 20443 - 20453. [Abstract] [Full Text] [PDF]

				J. E. Vance Thematic Review Series: Glycerolipids. Phosphatidylserine and phosphatidylethanolamine in mammalian cells: two metabolically related aminophospholipids J. Lipid Res., July 1, 2008; 49(7): 1377 - 1387. [Abstract] [Full Text] [PDF]

				C. Gkogkas, S. Middleton, A. M. Kremer, C. Wardrope, M. Hannah, T. H. Gillingwater, and P. Skehel VAPB interacts with and modulates the activity of ATF6 Hum. Mol. Genet., June 1, 2008; 17(11): 1517 - 1526. [Abstract] [Full Text] [PDF]

				S. Saito, H. Matsui, M. Kawano, K. Kumagai, N. Tomishige, K. Hanada, S. Echigo, S. Tamura, and T. Kobayashi Protein Phosphatase 2C{epsilon} Is an Endoplasmic Reticulum Integral Membrane Protein That Dephosphorylates the Ceramide Transport Protein CERT to Enhance Its Association with Organelle Membranes J. Biol. Chem., March 7, 2008; 283(10): 6584 - 6593. [Abstract] [Full Text] [PDF]

				G. M. Carman and S. A. Henry Phosphatidic Acid Plays a Central Role in the Transcriptional Regulation of Glycerophospholipid Synthesis in Saccharomyces cerevisiae J. Biol. Chem., December 28, 2007; 282(52): 37293 - 37297. [Full Text] [PDF]

				G.-S. Han, S. Siniossoglou, and G. M. Carman The Cellular Functions of the Yeast Lipin Homolog Pah1p Are Dependent on Its Phosphatidate Phosphatase Activity J. Biol. Chem., December 21, 2007; 282(51): 37026 - 37035. [Abstract] [Full Text] [PDF]

				C. J.R. Loewen, B. P. Young, S. Tavassoli, and T. P. Levine Inheritance of cortical ER in yeast is required for normal septin organization J. Cell Biol., November 5, 2007; 179(3): 467 - 483. [Abstract] [Full Text] [PDF]

				E. Teuling, S. Ahmed, E. Haasdijk, J. Demmers, M. O. Steinmetz, A. Akhmanova, D. Jaarsma, and C. C. Hoogenraad Motor Neuron Disease-Associated Mutant Vesicle-Associated Membrane Protein-Associated Protein (VAP) B Recruits Wild-Type VAPs into Endoplasmic Reticulum-Derived Tubular Aggregates J. Neurosci., September 5, 2007; 27(36): 9801 - 9815. [Abstract] [Full Text] [PDF]

				L. C. Hancock, R. P. Behta, and J. M. Lopes Genomic Analysis of the Opi- Phenotype Genetics, June 1, 2006; 173(2): 621 - 634. [Abstract] [Full Text] [PDF]

				R. J. Perry and N. D. Ridgway Oxysterol-binding Protein and Vesicle-associated Membrane Protein-associated Protein Are Required for Sterol-dependent Activation of the Ceramide Transport Protein Mol. Biol. Cell, June 1, 2006; 17(6): 2604 - 2616. [Abstract] [Full Text] [PDF]

				I. Hamamoto, Y. Nishimura, T. Okamoto, H. Aizaki, M. Liu, Y. Mori, T. Abe, T. Suzuki, M. M. C. Lai, T. Miyamura, et al. Human VAP-B Is Involved in Hepatitis C Virus Replication through Interaction with NS5A and NS5B J. Virol., November 1, 2005; 79(21): 13473 - 13482. [Abstract] [Full Text] [PDF]