The yeast cyclin-dependent kinase inhibitor Sic1 and mammalian p27Kip1 are functional homologues with a structurally conserved inhibitory domain

Abstract

In Saccharomyces cerevisiae, Sic1, an inhibitor of Cdk (cyclin-dependent kinase), blocks the activity of S-Cdk1 (Cdk1/Clb5,6) kinase that is required for DNA replication. Deletion of Sic1 causes premature DNA replication from fewer origins, extension of the S phase and inefficient separation of sister chromatids during anaphase. Despite the well-documented relevance of Sic1 inhibition of S-Cdk1 for cell cycle control and genome instability, the molecular mechanism by which Sic1 inhibits S-Cdk1 activity remains obscure. In this paper, we show that Sic1 is functionally and structurally related to the mammalian Cki (Cdk inhibitor) p27Kip1 of the Kip/Cip family. A molecular model of the inhibitory domain of Sic1 bound to the Cdk2–cyclin A complex suggested that the yeast inhibitor might productively interface with the mammalian Cdk2–cyclin A complex. Consistent with this, Sic1 is able to bind to, and strongly inhibit the kinase activity of, the Cdk2–cyclin A complex. In addition, comparison of the different inhibitory patterns obtained using histone H1 or GST (glutathione S-transferase)–pRb (retinoblastoma protein) fusion protein as substrate (the latter of which recognizes both the docking site and the catalytic site of Cdk2–cyclin A) offers interesting suggestions for the inhibitory mechanism of Sic1. Finally, overexpression of the KIP1 gene in vivo in Saccharomyces cerevisiae, like overexpression of the related SIC1 gene, rescues the cell cycle-related phenotype of a sic1Δ strain. Taken together, these findings strongly indicate that budding yeast Sic1 and mammalian p27Kip1 are functional homologues with a structurally conserved inhibitory domain.