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Volume 270, Number 12, Issue of March 24, 1995 pp. 6843-6855
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.
Maternal Xenopus Cdk2-Cyclin E Complexes Function during Meiotic and Early Embryonic Cell Cycles That Lack a G Phase (*)

(Received for publication, August 16, 1994; and in revised form, January 13, 1995)

Rachel E. Rempel (§) Susan B. Sleight James L. Maller (¶)

From the Howard Hughes Medical Institute and Department of Pharmacology, University of Colorado School of Medicine, Denver, Colorado 80262


ABSTRACT

Earlier work demonstrated that cyclins A1, B1, and B2 are not associated with Cdk2 from unfertilized Xenopus eggs. As a potential Cdk2 partner during meiosis, a cyclin E homolog was cloned from a Xenopus oocyte cDNA library and found to be 60% identical at the amino acid level to human cyclin E. Cyclin E1 protein was detected in resting oocytes, and the level increased severalfold in meiosis II, concomitant with the appearance of forms with decreased electrophoretic mobility. During oocyte maturation, the patterns of cyclin E1-associated kinase activity and Cdk2 activity were identical, with activity low until after germinal vesicle breakdown, peaking during meiosis II. Cyclin E1 complexes immunoprecipitated from unfertilized Xenopus eggs contained Cdk2 but not Cdc2. In cycling egg extracts Cdk2-cyclin E1-associated kinase activity oscillated, but the level of cyclin E1 protein and its association with Cdk2 did not vary appreciably; complex activity appeared to be regulated neither by the synthesis and destruction of the cyclin subunit nor by association/disassociation of the two subunits. During the early cleavage divisions in embryos, cyclin E1 and Cdk2 remained associated. The data indicate that the Cdk2-cyclin E complex functions during meiotic and embryonic cell cycles in addition to performing its established role during G(1) in somatic cells.


FOOTNOTES

*
This work was supported by Grant GM26743 from the National Institutes of Health. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked ``advertisement'' in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank(TM)/EMBL Data Bank with accession number(s) L23857[GenBank].

§
Associate of the Howard Hughes Medical Institute.

Investigator of the Howard Hughes Medical Institute. To whom correspondence should be addressed.

(^1)
The abbreviations used are: MPF, maturation-promoting factor; CSF, cytostatic factor; EB, extraction buffer; Pipes, 1,4-piperazinediethanesulfonic acid; DTT, dithiothreitol; PBS, phosphate-buffered saline: GVBD, germinal vesicle breakdown.

(^2)
Dr. T. Kishimoto, Tokyo Institute of Technology, Tokyo, personal communication.

(^3)
Dr. M. J. Cockerill and Dr. T. Hunt, Imperial Cancer Research Fund, South Mimms, United Kingdom, personal communication.

(^4)
Dr. J. Blow, Imperial Cancer Research Fund, South Mimms, United Kingdom, personal communication.

(^5)
R. Hartley, R. Rempel, and J. Maller, manuscript in preparation.

(^6)
Dr. T. Hunt, Imperial Cancer Research Fund, South Mimms, United Kingdom, personal communication.

(^7)
J. Su, E. Erikson, and J. Maller, unpublished results.


ACKNOWLEDGEMENTS

We thank Brad Lattes and Andrea Lewellyn for technical assistance, Olivier Haccard for help with computerized sequence analysis, Rebecca Hartley for assistance with embryo staging, and Eleanor Erikson for a critical reading of the manuscript.


©1995 by The American Society for Biochemistry and Molecular Biology, Inc.


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