A dominant negative RAS-specific guanine nucleotide exchange factor reverses neoplastic phenotype in K-ras transformed mouse fibroblasts

Paola Bossù, Marco Vanoni, Valeria Wanke, Maria Paola Cesaroni, Franco Tropea, Gabriella Melillo, Cinzia Asti, Stefano Porzio, Paolo Ruggiero, Vito Di Cioccio, Giovanni Maurizi, Annibale Ciabini, Lilia Alberghina

Research output: Contribution to journalArticlepeer-review

Abstract

Ras proteins are small GTPases playing a pivotal role in cell proliferation and differentiation. Their activation state depends on the competing action of GTPase Activating Proteins (GAP) and Guanine nucleotide Exchange Factors (GEF). A tryptophan residue (Trp1056 in CDC25(Mm)-GEF), conserved in all ras-specific GEFs identified so far has been previously shown to be essential for GEF activity. Its substitution with glutamic acid results in a catalytically inactive mutant, which is able to efficiently displace wild-type GEF from p21(ras) and to originate a stable ras/GEF binary complex due to the reduced affinity of the nucleotide-free ras GEF complex for the incoming nucleotide. We show here that this 'ras-sequestering property' can be utilized to attenuate ras signal transduction pathways in mouse fibroblasts transformed by oncogenic ras. In fact overexpression of the dominant negative GEF(W1056F) in stable transfected cells strongly reduces intracellular ras GTP levels in k-ras transformed fibroblasts. Accordingly, the transfected fibroblasts revert to wild-type phenotype on the basis of morphology, cell cycle and anchorage independent growth. The reversion of the transformed phenotype is accompanied by DNA endoreduplication. The possible use of dominant negative ras-specific GEFs as a tool to down-regulate tumor growth is discussed.

Original languageEnglish
Pages (from-to)2147-2154
Number of pages8
JournalOncogene
Volume19
Issue number17
Publication statusPublished - Apr 20 2000

Keywords

  • CDC25(Mm)
  • fos-luciferase
  • GEF
  • Oncogenes
  • Signal transduction therapy

ASJC Scopus subject areas

  • Molecular Biology
  • Cancer Research
  • Genetics

Fingerprint Dive into the research topics of 'A dominant negative RAS-specific guanine nucleotide exchange factor reverses neoplastic phenotype in K-ras transformed mouse fibroblasts'. Together they form a unique fingerprint.

Cite this