Related Experiment Videos
Inhibition of ras-induced germinal vesicle breakdown in Xenopus oocytes by rap-1B
M J Campa1, K J Chang, L Molina y Vedia
1Division of Cell Biology, Burroughs Wellcome Co., Research Triangle Park, NC 27709.
Abstract:
A cDNA clone (Krev-1) has recently been identified that possesses the ability to reverse the transformed phenotype when introduced into a K-ras-transformed NIH/3T3 cell line. The Krev-1 protein, also known as rap-1A, was found to share 50% homology with the ras proteins. The rap-1A protein has also been shown to block the interaction of ras with its GTPase activating protein in vitro, leading to speculation regarding its role in vivo. A closely related protein, rap-1B, has also been identified in platelets, human erythroleukemia cells, neutrophils, and aortic smooth muscle cells. Unlike rap-1A, rap-1B has been shown to be phosphorylated in platelets. Given the high degree of similarity between the amino acid sequences of rap-1A and rap-1B, we sought to investigate the effect of microinjected rap-1B on H-ras(Val12)-induced germinal vesicle breakdown in Xenopus laevis oocytes. In this assay system, equimolar concentrations of rap-1B were found to block germinal vesicle breakdown triggered by the oncogenic ras protein. However, in the presence of IGF-1, this inhibition was not observed. Moreover, rap-1B is readily phosphorylated in the oocytes.
Insights
The rap-1B protein inhibits oncogenic ras-induced cell cycle progression in Xenopus oocytes. This inhibition is reversed by IGF-1, suggesting rap-1B
Area of Science:
- Molecular Biology
- Cell Biology
- Oncology
Background:
- Krev-1 (rap-1A) reverses the transformed phenotype in K-ras-transformed cells.
- Rap-1A shares 50% homology with ras proteins and inhibits ras-GTPase activating protein interaction.
- Rap-1B, a related protein, is found in various human cell types and is phosphorylated in platelets.
Purpose of the Study:
- To investigate the effect of rap-1B on H-ras(Val12)-induced germinal vesicle breakdown in Xenopus laevis oocytes.
- To explore the potential role of rap-1B in ras-mediated signaling pathways.
Main Methods:
- Microinjection of rap-1B into Xenopus laevis oocytes.
- Induction of germinal vesicle breakdown using oncogenic H-ras(Val12).
- Assessment of rap-1B phosphorylation in oocytes.
Main Results:
- Equimolar concentrations of rap-1B blocked H-ras(Val12)-induced germinal vesicle breakdown.
- Inhibition by rap-1B was not observed in the presence of IGF-1.
- Rap-1B was readily phosphorylated in the oocytes.
Conclusions:
- Rap-1B acts as an inhibitor of oncogenic ras signaling in Xenopus oocytes.
- IGF-1 signaling may counteract the inhibitory effects of rap-1B.
- Rap-1B phosphorylation in oocytes suggests its involvement in intracellular signaling cascades.