Abstract
Previous work has shown that Suc1/Cks proteins can promote the hyperphosphorylation of primed Cdk1 substrates through the formation of ternary Cdk1-Cks-phosphosubstrate complexes. This raises the possibility that Cks proteins might be able to both facilitate and interfere with hyperphosphorylation through a mechanism analogous to the prozone effect in antigen-antibody interactions, with substoichiometric Cks promoting the formation of Cdk1-Cks-phosphosubstrate complexes and suprastoichiometric Cks instead promoting the formation of Cdk1-Cks and Cks-phosphosubstrate complexes. We tested this hypothesis through a combination of theory, proof-of-principle experiments with oligonucleotide annealing, and experiments on the interaction of Xenopus cyclin B1-Cdk1-Cks2 with Wee1A in vitro and in Xenopus extracts. Our findings help explain why both Cks under-expression and overexpression interfere with cell-cycle progression and provide insight into the regulation of the Cdk1 system.
Highlights
Mitotic cyclin-dependent kinase complexes consist of three proteins: the Cdk1 catalytic subunit, the allosteric activator cyclin B, and a third small protein referred to as Suc1 in S. pombe, Cks1 in S. cerevisiae, and Cks1 or 2 in vertebrates
Cks proteins are present throughout the eukaryotic kingdom and are well conserved ($50% amino acid identity between human Cks1 and its S. pombe and S. cerevisiae homologs)
Theory: The Prozone Effect in Ternary Complex Formation Bray and Lay (1997) examined a variety of protein oligomerization schemes through numerical simulations, looking for topologies that were capable of giving rise to a prozone effect
Summary
Mitotic cyclin-dependent kinase complexes consist of three proteins: the Cdk catalytic subunit, the allosteric activator cyclin B, and a third small protein referred to as Suc in S. pombe, Cks in S. cerevisiae, and Cks or 2 in vertebrates. Depleting Cks from extracts diminishes the Cdk1-dependent hyperphosphorylation of Cdc25C, Cdc (APC3), Myt, and Wee1A, and Cks promotes the hyperphosphorylation of these proteins in vitro (Patra and Dunphy, 1996, 1998; Patra et al, 1999). Adding excess Cks proteins to Xenopus diminishes substrate hyperphosphorylation (Dunphy and Newport, 1989). This raises the question of what exactly Cks proteins do, and why the phenotypes of Cks deficiency and excess are similar
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