Division of Intramural Research
Pulmonary Critical Care Medicine Branch

National Institutes of Health
10 Center Drive, Building 10 / Room 5N307
Bethesda, Maryland 20892-1434
Phone Number: 301-496-4554
Fax Number: 301-402-1610
Email Address: vaughanm@nhlbi.nih.gov

Research Interest(s):

Molecular & Cellular Biology
Structural Biology

Research Description:

ARF GTPases: Mechanisms of Signaling and Regulatory Functions

Our research deals with the ARF (ADP-ribosylation factor) GTP-binding proteins that are critical in vesicular transport and activate phospholipase D as well as phosphatidylinositol 5-kinase. Our goals are to define the physiological roles of specific ARFs and elucidate mechanisms of their molecular interactions. ARF function requires alternation between GTP-bound active and GDP-bound inactive forms. Activation requires a guanine nucleotide-exchange protein or GEP. There are several families of GEPs that differ widely in structure and regulatory properties. We use tools of cell and molecular biology and protein chemistry to elucidate mechanisms by which GEPs are regulated and themselves regulate ARF and other molecules. At present, the ~ 50-kDa GEP family of four cytohesins that are involved in integrin-mediated cell adhesion are a particular focus of our questions about cell-specific expression, control of alternative mRNA splicing to yield proteins with different phospholipid-binding specificities, and the relationship of ARF activation to cytostructural changes and cell adhesion. These phenomena presumably reflect scaffolding or anchoring interactions of cytohesins that integrate vital mechanical and signaling functions. We had earlier purified and cloned two ~200-kDa GEPs that are inhibited by brefeldin A, a drug that reversibly blocks protein secretion and causes apparent disintegration of the Golgi. These similar molecules are not only larger than the cytohesins but also differ considerably in overall structure. Our current studies of their interactions are revealing the previously unrecognized intersection of some ARF functions with other signalling pathways. Continuing studies of ARD1, a protein that we identified and cloned several years ago are also important. ARD1 contains an N-terminal GAP domain (p5) that accelerates GTP hydrolysis by the C-terminal ARF domain (p3) and inactivates ARD1. We described the effects of numerous mutations in separately synthesized p3 and p5 on GTPase activity and protein-protein interactions, as well as sequence motifs that signal lysosomal (in p5) or Golgi (in p3) localization and residues responsible for ARD interaction with and activation by cytohesin-1. Our ARD1 "knock-out" project is now beginning to provide information regarding its physiological role(s).

Selected Publications:

Structural Basis for the Inhibitory Effect of Brefeldin A on Guanine Nucleotide-exchange Proteins for ADP-ribosylation Factors. M Sata, J Moss, M Vaughan. Proceedings of the National Academy of Science U.S.A. 96:2752-2757(1999). PubMed

Brefeldin A Inhibited Activity of the Sec7 Domain of p200, a Mammalian Guanine Nucleotide-exchange Protein for ADP-ribosylation Factors. N Morinaga, R Adamik, J Moss, M Vaughan. Journal of Biological Chemistry 274:17417-17423(1999). PubMed

Similarities in Function and Gene Structure of Cytohesin-4 and Cytohesin-1, Guanine Nucleotide-exchange Proteins for ADP-ribosylation Factors. M Ogasawara, S-C Kim, R Adamik, A Togawa, VJ Ferrans, K Takeda, M Kirby, J Moss, M Vaughan. Journal of Biological Chemistry 275:3221-3230(2000). PubMed

Identification of Lyosomal and Golgi Localization Signals in GAP and ARF Domains of ARF Domain Protein 1. N Vitale, VJ Ferrans, J Moss, M Vaughan. Molecular Cellular Biology 20:7342-7352(2000). PubMed

Cytohesin-1 in 2001. J Moss, M Vaughan. Arch. Biochem. Biophys 397:156-61 (2002). PubMed

Last updated August 29, 2002

Principal Investigators

• Joel Moss
• Martha Vaughan
• Vincent Manganiello
• Bernadette Gochuico
• Stewart Levine