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Release from G1 growth arrest by transforming growth factor beta 1 requires cellular ras activity
P H Howe1, S F Dobrowolski, K B Reddy
1Department of Cell Biology, Cleveland Clinic Research Institute, Ohio 44195.
Abstract:
Transforming growth factor beta 1 (TGF beta 1) is a potent inhibitor of epithelial cell growth, although the mechanism of growth inhibition remains unknown. We report here a critical relationship between cellular p21ras activity and TGF beta 1 action. Microinjection of oncogenic Ha-ras protein into TGF beta 1-arrested mink lung epithelial cells overcomes TGF beta 1 growth inhibition and allows progression into S phase. Cells released from TGF beta 1 inhibition following microinjection with anti-p21ras antibody, on the other hand, remain TGF beta 1-arrested and do not enter S phase, indicating a requirement for p21ras activity. These biological data are substantiated biochemically in that TGF beta 1 is shown to decrease the activation state of endogenous p21ras, as measured by the level of GTP-bound p21ras. In addition, the phosphorylation and kinase activity of mitogen-activated protein kinase, which depends upon cellular ras activity, is elevated in cells which have been released from growth arrest by TGF beta 1. Together these data demonstrate the involvement of p21ras activity in TGF beta 1-induced growth inhibition and suggest that the inhibitor controls proliferation by modulating the activity of p21ras.
Insights
Transforming growth factor beta 1 (TGF beta 1) inhibits epithelial cell growth by reducing p21ras activity. Restoring p21ras function releases cells from TGF beta 1-induced growth arrest, revealing a key mechanism of proliferation control.
Area of Science:
- Cell Biology
- Molecular Biology
- Cancer Research
Background:
- Transforming growth factor beta 1 (TGF beta 1) is a known inhibitor of epithelial cell proliferation.
- The precise molecular mechanisms underlying TGF beta 1-mediated growth inhibition are not fully understood.
Purpose of the Study:
- To investigate the role of p21ras activity in TGF beta 1-induced epithelial cell growth inhibition.
- To elucidate the relationship between TGF beta 1 signaling and cellular ras pathway activation.
Main Methods:
- Microinjection of oncogenic Ha-ras protein and anti-p21ras antibody into TGF beta 1-treated mink lung epithelial cells.
- Biochemical assays to measure the activation state of endogenous p21ras (GTP-bound p21ras).
- Assessment of mitogen-activated protein kinase (MAPK) phosphorylation and kinase activity.
Main Results:
- Microinjection of Ha-ras protein reversed TGF beta 1-induced cell cycle arrest, allowing S phase entry.
- Inhibition of p21ras activity with an antibody prevented cells from exiting TGF beta 1-induced arrest.
- TGF beta 1 treatment decreased the activation state of endogenous p21ras.
- MAPK phosphorylation and kinase activity were elevated in cells released from TGF beta 1 arrest.
Conclusions:
- Cellular p21ras activity is critical for mediating TGF beta 1's inhibitory effects on epithelial cell proliferation.
- TGF beta 1 controls cell proliferation by modulating p21ras activity.
- These findings identify p21ras as a key downstream mediator in TGF beta 1 signaling pathways.