Interactions between Casein Kinase Iε (CKIε) and Two Substrates from Disparate Signaling Pathways Reveal Mechanisms for Substrate-Kinase Specificity

Abstract

BackgroundMembers of the Casein Kinase I (CKI) family of serine/threonine kinases regulate diverse biological pathways. The seven mammalian CKI isoforms contain a highly conserved kinase domain and divergent amino- and carboxy-termini. Although they share a preferred target recognition sequence and have overlapping expression patterns, individual isoforms often have specific substrates. In an effort to determine how substrates recognize differences between CKI isoforms, we have examined the interaction between CKIε and two substrates from different signaling pathways.Methodology/Principal FindingsCKIε, but not CKIα, binds to and phosphorylates two proteins: Period, a transcriptional regulator of the circadian rhythms pathway, and Disheveled, an activator of the planar cell polarity pathway. We use GST-pull-down assays data to show that two key residues in CKIα's kinase domain prevent Disheveled and Period from binding. We also show that the unique C-terminus of CKIε does not determine Dishevelled's and Period's preference for CKIε nor is it essential for binding, but instead plays an auxillary role in stabilizing the interactions of CKIε with its substrates. We demonstrate that autophosphorylation of CKIε's C-terminal tail prevents substrate binding, and use mass spectrometry and chemical crosslinking to reveal how a phosphorylation-dependent interaction between the C-terminal tail and the kinase domain prevents substrate phosphorylation and binding.Conclusions/SignificanceThe biochemical interactions between CKIε and Disheveled, Period, and its own C-terminus lead to models that explain CKIε's specificity and regulation.

A p120-catenin–CK1ε complex regulates Wnt signaling

“…p120-catenin interaction is relevant for CK1 action, since the cellular elimination of p120-catenin greatly decreases the activity of this protein kinase. It has been shown that auto-phosphorylation of CK1 C-tail prevents substrate access to the active site, therefore causing the auto-inhibition of the enzyme (Dahlberg et al, 2009). It is possible that binding to p120-catenin limits this autophosphorylation reaction.…”

Drosophila DBT Autophosphorylation of Its C-Terminal Domain Antagonized by SPAG and Involved in UV-Induced Apoptosis

“…3) (16,17). It has been proposed that the C terminus of mammalian CKIε undergoes a phosphorylation-dependent interaction with the kinase domain to prevent substrate binding and phosphorylation (18). Since the positions of serines and threonines in the DBT and CKI␦/ε termini are not well conserved, it is not expected that this phosphorylation-dependent regulatory interaction would be conserved.…”

“…It was first demonstrated that mammalian CKI␦ autophosphorylates its C-terminal domain to inhibit its activity (16), while a series of mutants with mutations in the C-terminal domain of CKIε were generated to identify specific phosphorylation residues that mediate inhibition (17). Others have proposed a model in which the phosphorylated C-terminal domain physically interacts with the catalytic domain to inhibit the kinase activity (18).…”

Role of CK1 in GSK3β-mediated phosphorylation and degradation of Snail

“…Taken together, our observations suggest that the Ser104 and Ser107 residues of Snail are phosphorylated by cellular CK1, but not by GSK3b, which serve as the priming sites for the subsequent phosphorylation of Snail by GSK3b. In addition, the characteristic kinase domain and/or C-terminal tail of CK1e (Dahlberg et al, 2009) might contribute to the CK1e-specific phosphorylation of Snail.…”