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Journal of Cell Science, Vol 112, Issue 9 1313-1324, Copyright © 1999 by Company of Biologists
JOURNAL ARTICLES |
SW James, KA Bullock, SE Gygax, BA Kraynack, RA Matura, JA MacLeod, KK McNeal, KA Prasauckas, PC Scacheri, HL Shenefiel, HM Tobin and SD Wade
Department of Biology, Gettysburg College, Gettysburg, PA 17325, USA. sjames@gettysburg.edu
The nimO predicted protein of Aspergillus nidulans is related structurally and functionally to Dbf4p, the regulatory subunit of Cdc7p kinase in budding yeast. nimOp and Dbf4p are most similar in their C-termini, which contain a PEST motif and a novel, short-looped Cys2-His2 zinc finger-like motif. DNA labelling and reciprocal shift assays using ts-lethal nimO18 mutants showed that nimO is required for initiation of DNA synthesis and for efficient progression through S phase. nimO18 mutants abrogated a cell cycle checkpoint linking S and M phases by segregating their unreplicated chromatin. This checkpoint defect did not interfere with other checkpoints monitoring spindle assembly and DNA damage (dimer lesions), but did prevent activation of a DNA replication checkpoint. The division of unreplicated chromatin was accelerated in cells lacking a component of the anaphase-promoting complex (bimEAPC1), consistent with the involvement of nimO and APC/C in separate checkpoint pathways. A nimO deletion conferred DNA synthesis and checkpoint defects similar to nimO18. Inducible nimO alleles lacking as many as 244 C-terminal amino acids supported hyphal growth, but not asexual development, when overexpressed in a ts-lethal nimO18 strain. However, the truncated alleles could not rescue a nimO deletion, indicating that the C terminus is essential and suggesting some type of interaction among nimO polypeptides.
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