title: Specific functional interaction of human cytohesin-1 and ADP-ribosylation factor domain protein (ARD1).
abstract: Activation of ADP-ribosylation factors (ARFs) is mediated by guanine nucleotide-exchange proteins, which accelerate conversion of inactive ARF-GDP to active ARF-GTP. ARF domain protein (ARD1), a 64-kDa GTPase with a C-terminal ADP-ribosylation factor domain, is localized to lysosomes and the Golgi apparatus. When ARD1 was used as bait to screen a human liver cDNA library using the yeast two-hybrid system, a cDNA for cytohesin-1, a approximately 50-kDa protein with ARF guanine nucleotide-exchange protein activity, was isolated. In this system, ARD1-GDP interacted well with cytohesin-1 but very poorly with cytohesin-2. In agreement, cytohesin-1, but not cytohesin-2, markedly accelerated [(35)S]guanosine 5'-3-O-(thio)triphosphate binding to ARD1. The effector region of the ARF domain of ARD1 appeared to be critical for the specific interaction with cytohesin-1. Replacement of single amino acids in the Sec7 domains of cytohesin-1 and -2 showed that residue 30 is critical for specificity. In transfected COS-7 cells, overexpressed ARD1 and cytohesin-1 were partially colocalized, as determined by confocal fluorescence microscopy. It was concluded that cytohesin-1 is likely to be involved in ARD1 activation, consistent with a role for ARD1 in the regulation of vesicular trafficking.
authors.affiliation: Pulmonary-Critical Care Medicine Branch and the Pathology Section, NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: N
authors.initials: N
authors.lastname: Vitale
authors.forename: G
authors.initials: G
authors.lastname: Pacheco-Rodriguez
authors.forename: V J
authors.initials: VJ
authors.lastname: Ferrans
authors.forename: W
authors.initials: W
authors.lastname: Riemenschneider
authors.forename: J
authors.initials: J
authors.lastname: Moss
authors.forename: M
authors.initials: M
authors.lastname: Vaughan
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D015815:Cell Adhesion Molecules / Q000737:chemistry* / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D015723:Gene Library; D020662:Guanine Nucleotide Exchange Factors; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection
chemical_list: D015815:Cell Adhesion Molecules; D020662:Guanine Nucleotide Exchange Factors; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C080681:TRIM23 protein, human; C100723:cytohesin-1; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins