Abstract

SummaryThe cyclin-dependent kinases (CDKs) are the major cell-cycle regulators that phosphorylate hundreds of substrates, controlling the onset of S phase and M phase [1, 2, 3]. However, the patterns of substrate phosphorylation increase are not uniform, as different substrates become phosphorylated at different times as cells proceed through the cell cycle [4, 5]. In fission yeast, the correct ordering of CDK substrate phosphorylation can be established by the activity of a single mitotic cyclin-CDK complex [6, 7]. Here, we investigate the substrate-docking region, the hydrophobic patch, on the fission yeast mitotic cyclin Cdc13 as a potential mechanism to correctly order CDK substrate phosphorylation. We show that the hydrophobic patch targets Cdc13 to the yeast centrosome equivalent, the spindle pole body (SPB), and disruption of this motif prevents both centrosomal localization of Cdc13 and the onset of mitosis but does not prevent S phase. CDK phosphorylation in mitosis is compromised for approximately half of all mitotic CDK substrates, with substrates affected generally being those that require the highest levels of CDK activity to become phosphorylated and those that are located at the SPB. Our experiments suggest that the hydrophobic patch of mitotic cyclins contributes to CDK substrate selection by directing the localization of Cdc13-CDK to centrosomes and that this localization of CDK contributes to the CDK substrate phosphorylation necessary to ensure proper entry into mitosis. Finally, we show that mutation of the hydrophobic patch prevents cyclin B1 localization to centrosomes in human cells, suggesting that this mechanism of cyclin-CDK spatial regulation may be conserved across eukaryotes.

Highlights

  • A further mechanism for generating differential cyclin-dependent kinases (CDKs) activity toward different substrates has been shown for the buddi

  • In the absence of Cdc13-CDK localization to the spindle pole body (SPB) during G2, roughly half of all detected mitotic CDK phosphosites are not efficiently phosphorylated. These correspond to substrates at the SPB and in the cytoplasm and are the substrates that require the highest levels of CDK activity to become phosphorylated

  • This suggests that correctly regulated Cdc13-CDK localization to the SPB during G2/M is required to reach the upper level of CDK activity at the SPB and in the cytoplasmic compartments necessary to phosphorylate these substrates and enter mitosis

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Summary

Graphical Abstract

Basu et al find that the substrate-docking region of cyclin B, the hydrophobic patch, governs this localization in both human cells and fission yeast. Without the cyclin B hydrophobic patch, CDK phosphorylation is impaired at mitosis but only in certain subcellular compartments. 2020, Current Biology 30, 883–892 March 9, 2020 a 2019 The Author(s).

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