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A characterization of cytostatic factor activity from Xenopus eggs and c-mos-transformed cells
I Daar1, R S Paules, G F Vande Woude
1ABL-Basic Research Program, NCI-Frederick Cancer Research and Development Center, Frederick, Maryland 21702.
Abstract:
In Xenopus oocytes, the mos proto-oncogene product is required during meiosis I for the activation of maturation promoting factor (MPF) and the subsequent breakdown of the germinal vesicle (GVBD). In addition, the mos product has been shown to be a candidate "initiator" of meiotic maturation and is an active component of cytostatic factor (CSF), an activity responsible for metaphase II arrest. Here we demonstrate that pp39mos is required throughout oocyte maturation. We found that in progesterone stimulated oocytes, depletion of mos RNA immediately before GVBD terminally decreased MPF. Likewise, oocytes depleted of mos RNA and induced to mature with crude MPF proceeded through GVBD but lacked the MPF activity required to arrest mature oocytes at metaphase II. Thus, during maturation the mos product is required, directly or indirectly, to sustain MPF activity. On the other hand, mouse NIH/3T3 cells transformed by the constitutive expression of pp39mosxc possessed CSF activity but lacked constitutive levels of MPF or its associated histone H1 kinase activity. Moreover, cytosols prepared from transformed NIH/3T3 cells or Xenopus eggs had similar levels of CSF activity, but pp39mos levels were greater than 40-fold higher in the transformed cell extract. These analyses show that maintenance of CSF during interphase does not result in the maintenance of MPF.
Insights
The mos proto-oncogene product is essential throughout Xenopus oocyte maturation for maintaining maturation promoting factor (MPF) activity. This ensures both germinal vesicle breakdown and metaphase II arrest, crucial for successful meiosis.
Area of Science:
- Cell Biology
- Molecular Biology
- Developmental Biology
Background:
- The mos proto-oncogene product is known to initiate meiotic maturation and is part of cytostatic factor (CSF), which causes metaphase II arrest in Xenopus oocytes.
- Its role in maintaining maturation promoting factor (MPF) activity and germinal vesicle breakdown (GVBD) during meiosis I has been established.
Purpose of the Study:
- To investigate the requirement of pp39mos throughout the entire process of oocyte maturation.
- To determine if mos is necessary for sustaining MPF activity and cytostatic factor (CSF) activity.
Main Methods:
- Depletion of mos RNA in progesterone-stimulated Xenopus oocytes before GVBD.
- Induction of maturation in mos-depleted oocytes using crude MPF.
- Analysis of MPF and CSF activity in mouse NIH/3T3 cells expressing pp39mosxc.
Main Results:
- Depletion of mos RNA before GVBD led to a terminal decrease in MPF activity.
- Oocytes depleted of mos matured through GVBD but failed to arrest at metaphase II due to lack of MPF activity.
- NIH/3T3 cells expressing pp39mosxc showed CSF activity but not constitutive MPF or H1 kinase activity, indicating CSF maintenance doesn't require high MPF.
Conclusions:
- The mos product is required throughout oocyte maturation to sustain MPF activity, directly or indirectly.
- Maintenance of CSF activity during interphase does not lead to sustained MPF activity.
- pp39mos is crucial for both initiating and maintaining the meiotic maturation process in Xenopus oocytes.