Physical and functional interactions between polo kinase and the spindle pole component Cut12 regulate mitotic commitment inS. pombe

Abstract

Commitment to mitosis is regulated by a protein kinase complex called MPF. MPF is inhibited by Wee1-related kinases and activated by Cdc25 phosphatase. MPF activation further boosts Cdc25 and represses Wee1. This feedback control probably involves polo kinase. A dominantcut12.s11mutation in theSchizosaccharomyces pombespindle pole body (SPB) component Cut12 both suppresses the conditional lethal mitotic commitment defect ofcdc25.22and promotes premature association of theS. pombepolo kinase, Plo1, with the SPB. We now show that Cut12 associated with Plo1 in two hybrid and immunoprecipitation assays. Plo1 function was required for recognition of the mitotic SPB by the phospho-specific antibody MPM-2. In vivo MPM-2 staining and in vitro kinase assays established that the loss-of-function mutation,cut12.1, reduced mitotic activation of Plo1, whereas the gain-of-function mutation,cut12.s11, promoted higher levels of Plo1 activity than were normally seen in interphase.cut12.s11could not promote mitotic commitment ofcdc25.22cells when Plo1 function was compromised. Expression of a constitutively activeplo1allele suppressed the mitotic commitment defect ofcdc25.22. These data suggest thatcut12.s11suppressescdc25.22by promoting Plo1 activity. Furthermore, the delayed mitotic commitment ofplo1.ts2cells suggests that Plo1 is an integral part of the core controls that modulate MPF activation inS. pombe.