Members of the Casein Kinase I (CKI) family of serine/threonine kinases regulate diverse biological pathways. The seven CKI isoforms contain a highly conserved kinase domain and divergent amino- (N-) and carboxyl- (C-) 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, I have examined the interaction between CKI&egr; and two substrates from divergent signaling pathways. CKI&egr;, but not CKIα, binds to and phosphorylates two proteins: Period (Per), a transcription factor in the circadian rhythms pathway, and Disheveled (Dsh), an activator of the planar cell polarity pathway. I have investigated the differences between CKI&egr; and CKIα in an attempt to understand what determines the interactions of two closely related and widely expressed kinases.

I present data showing that two key residues in CKIα's kinase domain prevent Dsh and Per from binding. I also show that the C-terminus of CKI&egr; does not determine Dsh and Per's preference for CKI&egr;, but that it is able to stabilize otherwise unfavorable interactions. Finally, I have investigated the importance of autophosphorylation of CKI&egr; with respect to the ability of substrates to bind to the kinase, as well as to change the conformational of the C-terminal quarter of the protein. The biochemical interactions between CKI&egr; and Dsh, Per, and its own C-terminus lead to models that help explain both CKI&egr;'s specificity and regulation in the cell.