title: NFBD1/MDC1 is phosphorylated by PLK1 and controls G2/M transition through the regulation of a TOPOIIα-mediated decatenation checkpoint.
abstract: Although it has been established that nuclear factor with BRCT domain 1/ mediator of the DNA damage checkpoint protein 1 (NFBD1/MDC1) is closely involved in DNA damage response, its possible contribution to the regulation of cell- cycle progression is unclear. In the present study, we have found for the first time that NFBD1 is phosphorylated by polo-like kinase 1 (PLK1) and has an important role in G2/M transition. Both NFBD1 and PLK1 are co-expressed in cellular nuclei throughout G2/M transition, and binding assays demonstrated direct interaction between NFBD1 and PLK1. Indeed, in vitro kinase reactions revealed that the PST domain of NFBD1 contains a potential amino acid sequence (845-DVTGEE-850) targeted by PLK1. Furthermore, enforced expression of GFP-PST but not GFP-PST(T847A) where threonine at 847 was substituted by alanine inhibited the phosphorylation levels of histone H3, suggesting a defect of M phase entry. Because PLK1 has been implicated in promoting the G2/M transition, we reasoned that overexpressed PST might serve as a pseudosubstrate for PLK1 and thus interfere with phosphorylation of endogenous PLK1 substrates. Interestingly, siRNA-mediated knockdown of NFBD1 resulted in early M phase entry and accelerated M phase progression, raising the possibility that NFBD1 is a PLK1 substrate for regulating the G2/M transition. Moreover, the constitutive active form of PLK1(T210D) overcame the ICRF-193-induced decatenation checkpoint and inhibited the interaction between NFBD1 and topoisomerase IIα, but kinase-deficient PLK1 did not. Based on these observations, we propose that PLK1-mediated phosphorylation of NFBD1 is involved in the regulation of G2/M transition by recovering a decatenation checkpoint.
authors.affiliation: Division of Biochemistry and Innovative Cancer Therapeutics, Chiba Cancer Center Research Institute, Chiba, Japan ; Division of Clinical Oncology Research, Shonan Kamakura General Hospital, Kanagawa, Japan.
authors.forename: Kiyohiro
authors.initials: K
authors.lastname: Ando
authors.affiliation: Laboratory of Anti-tumor Research, Chiba Cancer Center Research Institute, Chiba, Japan.
authors.forename: Toshinori
authors.initials: T
authors.lastname: Ozaki
authors.affiliation: Department of Experimental Pathology, Cancer Institute of the Japanese Foundation for Cancer Research (JFCR), Tokyo, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Hirota
authors.affiliation: Division of Biochemistry and Innovative Cancer Therapeutics, Chiba Cancer Center Research Institute, Chiba, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Nakagawara
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000951:Antigens, Neoplasm / Q000378:metabolism*; D002453:Cell Cycle / Q000502:physiology; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology*; D002460:Cell Line; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D016195:G2 Phase; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D006657:Histones / Q000378:metabolism; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D000097869:Polo-Like Kinase 1
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000951:Antigens, Neoplasm; D018797:Cell Cycle Proteins; D004250:DNA Topoisomerases, Type II; D004268:DNA-Binding Proteins; D006657:Histones; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D015534:Trans-Activators; D000097869:Polo-Like Kinase 1; C413163:MDC1 protein, human