Multisite phosphorylation by Cdk1 initiates delayed negative feedback to control mitotic transcription

Curr Biol. 2022 Jan 10;32(1):256-263.e4. doi: 10.1016/j.cub.2021.11.001. Epub 2021 Nov 23.


Cell-cycle progression is driven by the phosphorylation of cyclin-dependent kinase (Cdk) substrates.1-3 The order of substrate phosphorylation depends in part on the general rise in Cdk activity during the cell cycle,4-7 together with variations in substrate docking to sites on associated cyclin and Cks subunits.3,6,8-10 Many substrates are modified at multiple sites to provide more complex regulation.10-14 Here, we describe an elegant regulatory circuit based on multisite phosphorylation of Ndd1, a transcriptional co-activator of budding yeast genes required for mitotic progression.11,12 As cells enter mitosis, Ndd1 phosphorylation by Cdk1 is known to promote mitotic cyclin (CLB2) gene transcription, resulting in positive feedback.13-16 Consistent with these findings, we show that low Cdk1 activity promotes CLB2 expression at mitotic entry. We also find, however, that when high Cdk1 activity accumulates in a mitotic arrest, CLB2 expression is inhibited. Inhibition is accompanied by Ndd1 degradation, and we present evidence that degradation is triggered by multisite Ndd1 phosphorylation by high mitotic Cdk1-Clb2 activity. Complete Ndd1 phosphorylation by Clb2-Cdk1-Cks1 requires the phosphothreonine-binding site of Cks1, as well as a recently identified phosphate-binding pocket on the cyclin Clb2.17 We therefore propose that initial phosphorylation by Cdk1 primes Ndd1 for delayed secondary phosphorylation at suboptimal sites that promote degradation. Together, our results suggest that rising levels of mitotic Cdk1 activity act at multiple phosphorylation sites on Ndd1, first triggering rapid positive feedback and then promoting delayed negative feedback, resulting in a pulse of mitotic gene expression.

Keywords: Cdk; Cks1; Clb2; Ndd1; cell cycle; cyclin gene expression; multisite phosphorylation; phosphodegron.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • CDC2 Protein Kinase / genetics
  • CDC2 Protein Kinase / metabolism
  • Cell Cycle Proteins / metabolism
  • Cyclin B / genetics
  • Cyclin B / metabolism
  • Cyclins / genetics
  • Feedback
  • Mitosis
  • Phosphorylation
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins* / genetics
  • Saccharomyces cerevisiae Proteins* / metabolism
  • Transcription Factors / metabolism


  • Cell Cycle Proteins
  • Cyclin B
  • Cyclins
  • Saccharomyces cerevisiae Proteins
  • Transcription Factors
  • CDC2 Protein Kinase