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Induction of metaphase arrest in cleaving Xenopus embryos by MAP kinase
O Haccard1, B Sarcevic, A Lewellyn
1Howard Hughes Medical Institute, University of Colorado School of Medicine, Denver 80262.
Abstract:
The natural arrest of vertebrate unfertilized eggs in second meiotic metaphase results from the activity of cytostatic factor (CSF). The product of the c-mos(xe) proto-oncogene is thought to be a component of CSF and can induce metaphase arrest when injected into blastomeres of two-cell embryos. The c-Mos(xe) protein can directly activate the mitogen-activated protein kinase kinase (MAP kinase kinase) in vitro, leading to activation of MAP kinase. MAP kinase and c-Mos(xe) are active in unfertilized eggs and are rapidly inactivated after fertilization. Microinjection of thiophosphorylated MAP kinase into one blastomere of a two-cell embryo induced metaphase arrest similar to that induced by c-Mos(xe). However, only arrest with c-Mos(xe) was associated with activation of endogenous MAP kinase. These results indicate that active MAP kinase is a component of CSF in Xenopus and suggest that the CSF activity of c-Mos(xe) is mediated by MAP kinase.
Insights
Cytostatic factor (CSF) arrests unfertilized vertebrate eggs in meiosis. The c-Mos protein activates MAP kinase, which is essential for this metaphase arrest in Xenopus.
Area of Science:
- Cell Biology
- Developmental Biology
- Molecular Biology
Background:
- Unfertilized vertebrate eggs naturally arrest in second meiotic metaphase due to cytostatic factor (CSF) activity.
- The c-mos proto-oncogene product (c-Mos) is a suspected component of CSF and can induce metaphase arrest.
- Both c-Mos and MAP kinase are active in unfertilized eggs and inactivated upon fertilization.
Purpose of the Study:
- To investigate the role of MAP kinase in the cytostatic factor (CSF) activity of c-Mos.
- To determine if active MAP kinase is a component of CSF in Xenopus.
- To elucidate the mechanism by which c-Mos mediates metaphase arrest.
Main Methods:
- Injection of c-Mos(xe) protein into two-cell Xenopus embryos.
- In vitro activation assays of MAP kinase kinase (MAPKK) by c-Mos(xe).
- Microinjection of thiophosphorylated MAP kinase into Xenopus blastomeres.
Main Results:
- c-Mos(xe) injection induced metaphase arrest, mimicking CSF activity.
- c-Mos(xe) directly activated MAPKK, leading to MAP kinase activation in vitro.
- Microinjected MAP kinase induced metaphase arrest, but only c-Mos(xe) treatment activated endogenous MAP kinase.
Conclusions:
- Active MAP kinase is a crucial component of CSF in Xenopus.
- The CSF activity of c-Mos(xe) is mediated through the activation of MAP kinase.
- This study clarifies the molecular pathway regulating meiotic arrest in vertebrate eggs.