title: Vimentin phosphorylation and assembly are regulated by the small GTPase Rab7a.
abstract: Intermediate filaments are cytoskeletal elements important for cell architecture. Recently it has been discovered that intermediate filaments are highly dynamic and that they are fundamental for organelle positioning, transport and function thus being an important regulatory component of membrane traffic. We have identified, using the yeast two-hybrid system, vimentin, a class III intermediate filament protein, as a Rab7a interacting protein. Rab7a is a member of the Rab family of small GTPases and it controls vesicular membrane traffic to late endosomes and lysosomes. In addition, Rab7a is important for maturation of phagosomes and autophagic vacuoles. We confirmed the interaction in HeLa cells by co-immunoprecipitation and pull-down experiments, and established that the interaction is direct using bacterially expressed recombinant proteins. Immunofluorescence analysis on HeLa cells indicate that Rab7a-positive vesicles sometimes overlap with vimentin filaments. Overexpression of Rab7a causes an increase in vimentin phosphorylation at different sites and causes redistribution of vimentin in the soluble fraction. Consistently, Rab7a silencing causes an increase of vimentin present in the insoluble fraction (assembled). Also, expression of Charcot-Marie-Tooth 2B-causing Rab7a mutant proteins induces vimentin phosphorylation and increases the amount of vimentin in the soluble fraction. Thus, modulation of expression levels of Rab7a wt or expression of Rab7a mutant proteins changes the assembly of vimentin and its phosphorylation state indicating that Rab7a is important for the regulation of vimentin function.
authors: Department of Biological and Environmental Sciences and Technologies, University of Salento, Lecce, Italy.
authors: Laura
authors: L
authors: Cogli
authors: Cinzia
authors: C
authors: Progida
authors: Roberta
authors: R
authors: Bramato
authors: Cecilia
authors: C
authors: Bucci
mesh_terms: D015153:Blotting, Western; D011992:Endosomes; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014746:Vimentin / Q000378:metabolism*; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000090802:rab7 GTP-Binding Proteins
chemical_list: D050505:Mutant Proteins; D011994:Recombinant Proteins; D014746:Vimentin; D000090802:rab7 GTP-Binding Proteins; D020691:rab GTP-Binding Proteins