title: Functional characterization of Rab7 mutant proteins associated with Charcot-Marie-Tooth type 2B disease.
abstract: Charcot-Marie-Tooth (CMT) type 2 neuropathies are a group of autosomal-dominant axonal disorders genetically and clinically heterogeneous. In particular, CMT type 2B (CMT2B) neuropathies are characterized by severe sensory loss, often complicated by infections, arthropathy, and amputations. Recently, four missense mutations in the small GTPase Rab7 associated with the Charcot-Marie Tooth type 2B phenotype have been identified. These mutations target highly conserved amino acid residues. However, nothing is known about whether and how these mutations affect Rab7 function. We investigated the biochemical and functional properties of three of the mutant proteins. Interestingly, all three proteins exhibited higher nucleotide exchange rates and hydrolyzed GTP slower than the wild-type protein. In addition, whereas 23% of overexpressed wild-type Rab7 was GTP bound in HeLa cells, the large majority of the mutant proteins (82-89%) were in the GTP-bound form, consistent with the data on GTP hydrolysis and exchange rates. The CMT2B-associated Rab7 proteins were also able to bind the Rab7 effector RILP (Rab-interacting lysosomal protein) and to rescue Rab7 function after silencing. Altogether, these data demonstrate that all tested CMT2B-associated Rab7 mutations are mechanistically similar, suggesting that activated forms of the Rab7 are responsible for CMT2B disease.
authors.affiliation: Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento, 73100 Lecce, Italy.
authors.forename: Maria Rita
authors.initials: MR
authors.lastname: Spinosa
authors.forename: Cinzia
authors.initials: C
authors.lastname: Progida
authors.forename: Azzurra
authors.initials: A
authors.lastname: De Luca
authors.forename: Anna Maria Rosaria
authors.initials: AM
authors.lastname: Colucci
authors.forename: Pietro
authors.initials: P
authors.lastname: Alifano
authors.forename: Cecilia
authors.initials: C
authors.lastname: Bucci
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000595:Amino Acid Sequence; D002607:Charcot-Marie-Tooth Disease / Q000235:genetics*; D004705:Endocytosis; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D050505:Mutant Proteins / Q000378:metabolism; D020125:Mutation, Missense; D020691:rab GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000090802:rab7 GTP-Binding Proteins
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D050505:Mutant Proteins; C423317:RILP protein, human; D000090802:rab7 GTP-Binding Proteins; C000716463:rab7 GTP-binding proteins, human; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D020691:rab GTP-Binding Proteins