Proteolysis and Tyrosine Phosphorylation of p34 cdc2 /Cyclin B

doi:10.1074/jbc.272.52.33384

Proteolysis and Tyrosine Phosphorylation of p34^cdc2/Cyclin B

THE ROLE OF MCM2 AND INITIATION OF DNA REPLICATION TO ALLOW TYROSINE PHOSPHORYLATION OF p34^cdc2*

From the ^‡Henry Hood Research Program, Weis Center for Research, Pennsylvania State University College of Medicine, Danville, Pennsylvania 17822 and the ^§Department of Biology, Gettysburg College, Gettysburg, Pennsylvania 17325

Abstract

Previously, it has been shown thatAspergillus cells lacking the function of nimQand the anaphase-promoting complex (APC) componentbimE ^APC1 enter mitosis without replicating DNA. Here nimQ is shown to encode an MCM2 homologue. Although mutation of nimQ ^MCM2 inhibits initiation of DNA replication, a few cells do enter mitosis. Cells arrested at G₁/S by lack ofnimQ ^MCM2 contain p34^cdc2/cyclin B, but p34^cdc2 remains tyrosine dephosphorylated, even after DNA damage. However, arrest of DNA replication using hydroxyurea followed by inactivation of nimQ ^MCM2 andbimE ^APC1 does not abrogate the S phase arrest checkpoint over mitosis. nimQ ^MCM2, likely via initiation of DNA replication, is therefore required to trigger tyrosine phosphorylation of p34^cdc2 during the G₁to S transition, which may occur by inactivation ofnimT ^cdc25. Cells lacking bothnimQ ^MCM2 and bimE ^APC1are deficient in the S phase arrest checkpoint over mitosis because they lack both tyrosine phosphorylation of p34^cdc2 and the function of bimE ^APC1. Initiation of DNA replication, which requires nimQ ^MCM2, is apparently critical to switch mitotic regulation from the APC to include tyrosine phosphorylation of p34^cdc2 at G₁/S. We also show that cells arrested at G₁/S due to lack of nimQ ^MCM2 continue to replicate spindle pole bodies in the absence of DNA replication and can undergo anaphase in the absence of APC function.

Footnotes

↵* This work was supported by a grant to Gettysburg College from the Howard Hughes Medical Institute, by grants from Gettysburg College (to S. W. J.), and by Grant GM42564 from the National Institutes of Health (to S. A. O.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be 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™/EMBL Data Bank with accession number(s) AF014813.
↵¶ To whom correspondence should be addressed. Tel.: 717-271-6677; Fax: 717-271-6701; E-mail: sao{at}psghs.edu.
↵1 The abbreviations used are: MPF, mitosis- or maturation-promoting factor; APC, anaphase-promoting complex; SPB, spindle pole body; DAPI, 4′6-diamidino-2-phenylindole; HU, hydroxyurea; kb, kilobase pair(s); MMS, methyl methanesulfonate.
↵2 G. E. Crawford, A. N. Wexler, and S. W. James, unpublished data.
- Received September 23, 1997.
- Revision received October 21, 1997.