title: Characterization and properties of dominant-negative mutants of the ras-specific guanine nucleotide exchange factor CDC25(Mm).
abstract: Ras proteins are small GTPases playing a pivotal role in cell proliferation and differentiation. Their activation depends on the competing action of GTPase activating proteins and guanine nucleotide exchange factors (GEF). The properties of two dominant-negative mutants within the catalytic domains of the ras-specific GEF, CDC25(Mm), are described. In vitro, the mutant GEF(W1056E) and GEF(T1184E) proteins are catalytically inactive, are able to efficiently displace wild-type GEF from p21(ras), and strongly reduce affinity of the nucleotide-free ras x GEF complex for the incoming nucleotide, thus resulting in the formation of a stable ras.GEF binary complex. Consistent with their in vitro properties, the two mutant GEFs bring about a dramatic reduction in ras-dependent fos-luciferase activity in mouse fibroblasts. The stable ectopic expression of the GEF(W1056E) mutant in smooth muscle cells effectively reduced growth rate and DNA synthesis with no detectable morphological changes.
authors.affiliation: Dipartimento di Biotecnologie e Bioscienze, Università degli Studi di Milano-Bicocca, Piazza della Scienza, 2 20126 Milano, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Vanoni
authors.forename: R
authors.initials: R
authors.lastname: Bertini
authors.forename: E
authors.initials: E
authors.lastname: Sacco
authors.forename: L
authors.initials: L
authors.lastname: Fontanella
authors.forename: M
authors.initials: M
authors.lastname: Rieppi
authors.forename: S
authors.initials: S
authors.lastname: Colombo
authors.forename: E
authors.initials: E
authors.lastname: Martegani
authors.forename: V
authors.initials: V
authors.lastname: Carrera
authors.forename: A
authors.initials: A
authors.lastname: Moroni
authors.forename: C
authors.initials: C
authors.lastname: Bizzarri
authors.forename: V
authors.initials: V
authors.lastname: Sabbatini
authors.forename: M
authors.initials: M
authors.lastname: Cattozzo
authors.forename: A
authors.initials: A
authors.lastname: Colagrande
authors.forename: L
authors.initials: L
authors.lastname: Alberghina
mesh_terms: D000818:Animals; D002460:Cell Line; D051379:Mice; D009131:Muscle, Smooth, Vascular / Q000502:physiology*; D009154:Mutation; D051381:Rats; D015398:Signal Transduction / Q000235:genetics; D020666:ras-GRF1 / Q000235:genetics* / Q000378:metabolism
chemical_list: D020666:ras-GRF1