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PMID: 7744959 Published · ppublish English Journal Article

R-Ras promotes apoptosis caused by growth factor deprivation via a Bcl-2 suppressible mechanism.

The Journal of cell biology ·Vol. 129 ·No. 4 ·1995-05-00 ·Pages 1103-14

Wang HG, Millan JA, Cox AD, Der CJ, Rapp UR, Beck T, Zha H, Reed JC

Abstract

The Bcl-2 protein is an important regulator of programmed cell death, but the biochemical mechanism by which this protein prevents apoptosis remains enigmatic. Recently, Bcl-2 has been reported to physically interact with a member of the Ras superfamily of small GTPases, p23-R-Ras. To examine the functional significance of R-Ras for regulation of cell death pathways, the IL-3-dependent cells 32D.3 and FL5.12 were stably transfected with expression plasmids encoding an activated form (38 Glycine-->Valine) of R-Ras protein. R-Ras(38V)-producing 32D.3 and FL5.12 cells experienced increased rates of apoptotic cell death relative to control transfected cells when deprived of IL-3. Analysis of several independent clones of transfected 32D.3 cells revealed a correlation between higher levels of R-Ras protein and faster rates of cell death upon withdrawal of IL-3 from cultures. 32D.3 cells cotransfected with R-Ras(38V) and Bcl-2 exhibited prolonged cell survival in the absence of IL-3, equivalent to 32D.3 cells transfected with Bcl-2 expression plasmids alone. R-Ras(38V) also increased rates of cell death in serum-deprived NIH-3T3 cells, and Bcl-2 again abrogated most of this effect. The ratio of GTP and GDP bound to R-Ras(38V) was not significantly different in control 32D.3 cells vs those that overexpressed Bcl-2, indicating that Bcl-2 does not abrogate R-Ras-mediated effects on cell death by altering R-Ras GDP/GTP regulation. Moreover, purified Bcl-2 protein had no effect on the GTPase activity of recombinant wild-type R-Ras in vitro. When expressed in Sf9 cells using recombinant baculoviruses, R-Ras(38V) bound to and induced activation of Raf-1 kinase irrespective of whether Bcl-2 was coproduced in these cells, suggesting that Bcl-2 does not nullify R-Ras effects by interfering with R-Ras-mediated activation of Raf-1 kinase. Taken together, these findings suggest that R-Ras enhances the activity of a cell death pathway in growth factor-deprived cells and imply that Bcl-2 acts downstream of R-Ras to promote cell survival.

MeSH Terms
3T3 Cells Animals Apoptosis/genetics,physiology Cells, Cultured Clone Cells Culture Media, Serum-Free GTP Phosphohydrolases/metabolism GTP-Binding Proteins/metabolism Growth Substances/deficiency Guanosine Diphosphate/metabolism Guanosine Triphosphate/metabolism Mice Precipitin Tests Protein Binding Protein Serine-Threonine Kinases/metabolism Proto-Oncogene Proteins/genetics,immunology,metabolism Proto-Oncogene Proteins c-bcl-2 Proto-Oncogene Proteins c-raf Suppression, Genetic Transfection bcl-2-Associated X Protein ras Proteins/metabolism
Chemicals
Culture Media, Serum-Free Growth Substances Proto-Oncogene Proteins Proto-Oncogene Proteins c-bcl-2 bcl-2-Associated X Protein Guanosine Diphosphate Guanosine Triphosphate Protein Serine-Threonine Kinases Proto-Oncogene Proteins c-raf GTP Phosphohydrolases GTP-Binding Proteins Rras protein, mouse ras Proteins
Authors & Affiliations
8 authors, click to expand affiliations / ORCID
Wang H G
La Jolla Cancer Research Foundation, California 92037, USA.
Millan J A
Cox A D
Der C J
Rapp U R
Beck T
Zha H
Reed J C
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.129.4.1103
Published
1995-05-00
Pages
1103-14
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2120494
Subset
IM
External Links
PubMed source DOI: 10.1083/jcb.129.4.1103 PMC Full Text View journal details
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