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Oncogenic Ras blocks cell cycle progression and inhibits p34cdc2 kinase in activated Xenopus egg extracts
1Department of Surgery, University of Kentucky Medical Center, Lexington 40536.
Abstract:
The effect of purified, bacterially expressed human RasH proteins on embryonic cell cycle progression in activated Xenopus egg extracts was studied. Bacterially expressed human oncogenic RasH protein is able to block the progression of the Xenopus embryonic cell cycle into M-phase. In contrast, the corresponding normal human Ras protein is relatively ineffective when assayed in a like manner. The observed arresting activity can be blocked by the addition of Y13-259 anti-Ras monoclonal antibody but not by nonspecific IgG. Oncogenic Ras also induces unique morphological changes in the reconstituted nuclei; the nuclei appear to be enlarged, and the chromatin partially condenses into fiber-like structures. This induced arrest is associated with suppression of p34cdc2 kinase activity, indicating that the oncogenic Ras protein induces the arrest by inactivating p34cdc2. This inactivation by oncogenic Ras protein does not result from inhibition of the synthesis of cyclin B or of the binding of the newly synthesized cyclin B to the p34cdc2.
Insights
Oncogenic RasH protein halts the Xenopus embryonic cell cycle at M-phase, unlike normal RasH. This effect, blocked by anti-Ras antibodies, involves p34cdc2 inactivation.
Area of Science:
- Cell Biology
- Molecular Biology
- Developmental Biology
Background:
- Ras proteins are key regulators of cell signaling.
- Aberrant Ras signaling is implicated in cancer development.
- Understanding Ras function in cell cycle control is crucial.
Purpose of the Study:
- To investigate the effect of human RasH proteins on the Xenopus embryonic cell cycle.
- To determine if oncogenic RasH has a different effect compared to normal RasH.
- To elucidate the molecular mechanism underlying Ras-induced cell cycle arrest.
Main Methods:
- Utilized activated Xenopus egg extracts for cell cycle studies.
- Employed bacterially expressed purified human RasH proteins (oncogenic and normal).
- Assessed cell cycle progression, nuclear morphology, and p34cdc2 kinase activity.
Main Results:
- Oncogenic RasH protein blocked Xenopus embryonic cell cycle progression into M-phase.
- Normal human Ras protein showed minimal effect on cell cycle progression.
- The cell cycle arrest was specifically inhibited by an anti-Ras monoclonal antibody.
- Oncogenic Ras induced nuclear enlargement and partial chromatin condensation.
- The arrest correlated with suppressed p34cdc2 kinase activity, independent of cyclin B synthesis or binding.
Conclusions:
- Oncogenic RasH protein actively inhibits embryonic cell cycle progression at M-phase.
- The mechanism involves the inactivation of p34cdc2 kinase by oncogenic RasH.
- These findings highlight a critical role for Ras proteins in cell cycle regulation and provide insights into oncogenic mechanisms.