Cdk1 phosphorylates SPAT-1/Bora to trigger PLK-1 activation and drive mitotic entry in C. elegans embryos.
Bottom Line: We further show that phospho-SPAT-1 activates PLK-1 by triggering phosphorylation on its activator T loop in vitro by Aurora A.Likewise, we show that phosphorylation of human Bora by Cdk1 promotes phosphorylation of human Plk1 by Aurora A, suggesting that this mechanism is conserved in humans.Our results suggest that CDK-1 activates PLK-1 via SPAT-1 phosphorylation to promote entry into mitosis.
Affiliation: Jacques Monod Institute, UMR7592; and Mass Spectrometry Facility, Jacques Monod Institute, UMR7592; Paris-Diderot University-Centre National de la Recherche Scientifique, 75013 Paris, France.Show MeSH
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Mentions: To identify the residues phosphorylated by CDK-1, we subjected SPAT-1, phosphorylated in vitro by human CyclinB/Cdk1, to tandem mass spectrometry (MS; liquid chromatography [LC]–MS/MS) analysis and identified 13 ((S/T)P) residues (S36, T57, T78, S119, S190, T229, S251, T328, S368, T456, T465, S504, and T518; Fig. 2, A and B; and Table S2). A nonphosphorylatable SPAT-113A mutant (with all S and T of the (S/T)P sites mutated to alanine) displayed a significantly dampened phosphorylation by CyclinB/Cdk1 (Fig. 2 C), indicating that these 13 residues are indeed the major CyclinB/Cdk1 phosphorylation sites. MS analysis of endogenous SPAT-1 or GFP::SPAT-1 immunoprecipitated from WT embryonic extracts confirmed that SPAT-1 is phosphorylated in vivo on at least 7 (S36, T57, S190, T229, S251, T328, and S504) out of the 13 residues that are phosphorylated by Cdk1 in vitro (Materials and methods; Fig. 2 B and Table S2).
Affiliation: Jacques Monod Institute, UMR7592; and Mass Spectrometry Facility, Jacques Monod Institute, UMR7592; Paris-Diderot University-Centre National de la Recherche Scientifique, 75013 Paris, France.