Biomedical subjects

S Stang

Publications and source records attributed to S Stang.

3 recordsLinked to original sources

PubMed1997

Interaction of activated Ras with Raf-1 alone may be sufficient for transformation of rat2 cells.

v-H-ras effector mutants have been assessed for transforming activity and for the ability of the encoded proteins to interact with Raf-1-, B-Raf-, byr2-, ralGDS-, and CDC25-encoded proteins in the yeast two-hybrid system. Transformation was assessed in rat2 cells as well as in a mutant cell line, rv68BUR, that affords a more sensitive transformation assay. Selected mutant Ras proteins were also examined for their ability to interact with an amino-terminal fragment of Raf-1 in vitro. Finally, possible cooperation between different v-H-ras effector mutants and between effector mutants and overexpressed Raf-1 was assessed. Ras transforming activity was shown to correlate best with the ability of the encoded protein to interact with Raf-1. No evidence for cooperation between v-H-ras effector mutants was found. Signaling through the Raf1-MEK-mitogen-activated protein kinase cascade may be the only effector pathway contributing to RAS transformation in these cells.

Animals↗

PubMed1996

ras effector loop mutations that dissociate p120GAP and neurofibromin interactions.

ras proteins are positively regulated by nucleotide exchange factors and negatively regulated by GTPase-activating proteins (GAPs). Two GAPs have been found in mammalian cells, p120GAP and neurofibromin, the product of the type 1 neurofibromatosis (NF1) gene. A library of substitutions in the effector loop region of ras in an Escherichia coli plasmid expression system was screened for c-Ha-ras species with altered GAP interactions. Several substitutions preferentially disrupted the interaction of ras with p120GAP as compared with the interaction with the recombinant GAP-related domain of neurofibromin (NF1-GRD). The most extreme example, Tyr32His, encoded a ras species that was unaffected by p120GAP but was stimulated normally by NF1-GRD. Tyr32His was weakly transforming in Rat2 cells. Tyr32His ras was primarily GDP-bound in quiescent Rat2 cells, although it rapidly associated with GTP after treatment of cells with epidermal growth factor. These results show that the NF1 product has less stringent requirements than p120GAP for ras effector domain structure and that negative regulation of ras can be achieved in rat fibroblasts by the product of NF1.

Animals↗

PubMed1995

RAS signalling is abnormal in a c-raf1 MEK1 double mutant.

A mutant rat cell clone that suppresses the transformation defects of RAS effector loop substitutions is heterozygous for mutations in c-raf1 and MEK1. The mutant cells can be transformed by many otherwise defective RAS effector mutants, including RAS genes with the effector regions of distantly related GTPases, even though the encoded RAS proteins do not interact with either the mutant or wild-type RAF in Saccharomyces cerevisiae. While the significance of the c-raf1 mutation is unclear, the MEK1 mutation increases MEK1 activity and leads to activation of mitogen-activated protein kinase. The mutant MEK1 is coupled to the epidermal growth factor pathway but exhibits decreased physical interaction with RAF. When overexpressed, the MEK1 mutation is transforming and causes hyperphosphorylation of RAF. Signalling from RAS to MEK1 may be mediated by something other than RAF alone, but signalling through MEK1 is probably sufficient for RAS transformation.

Amino Acid Sequence↗