title: Identification and characterisation of novel Mss4-binding Rab GTPases.
abstract: The Mss4 (mammalian suppressor of yeast Sec4) is an evolutionarily highly conserved protein and is expressed in all mammalian tissues. Although its precise biological function is still elusive, it has been shown to associate with a subset of secretory Rab proteins (Rab1b, Rab3a, Rab8a, Rab10) and to possess a rather low guanine nucleotide exchange factor (GEF) activity towards them in vitro (Rab1, Rab3a and Rab8a). By screening a human placenta cDNA library with Mss4 as bait, we identified several Rab GTPases (Rab12, Rab13 and Rab18) as novel Mss4-binding Rab proteins. Only exocytic but no endocytic Rab GTPases were found in our search. The binding of Mss4 to Rab proteins was confirmed by direct yeast two-hybrid interaction, by co-immunoprecipitation from lysates of mammalian cells, by immunofluorescence colocalisation as well as by direct in vitro binding studies. Analysis of Mss4 catalytic activity towards different Rab substrates confirmed that it is a somewhat inefficient GEF. These data, together with our mutational analysis of Mss4-Rab binding capacity, support the already proposed idea that Mss4 functions rather as a chaperone for exocytic Rab GTPases than as a GEF.
authors.affiliation: Institute of Molecular Virology, Münster University Hospital, Von Esmarch-Str. 56, D-48149 Münster, Germany.
authors.forename: Viktor
authors.initials: V
authors.lastname: Wixler
authors.forename: Ludmilla
authors.initials: L
authors.lastname: Wixler
authors.forename: Anika
authors.initials: A
authors.lastname: Altenfeld
authors.forename: Stephan
authors.initials: S
authors.lastname: Ludwig
authors.forename: Roger S
authors.initials: RS
authors.lastname: Goody
authors.forename: Aymelt
authors.initials: A
authors.lastname: Itzen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D005455:Fluorescent Antibody Technique; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D041681:NIH 3T3 Cells; D011485:Protein Binding; D020691:rab GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D018832:Molecular Chaperones; C489794:RABIF protein, human; D020691:rab GTP-Binding Proteins