Relationship between casein kinase I isoforms and a neurofilament-associated kinase

Purified neurofilaments (NFs) contain an associated kinase (NFAK) activity that phosphorylates selectively a subset of sites in the tail of NF-M and h as properties consistent with casein kinase I (CKI). Because CKI consists o f a family of as many as seven genes (alpha, beta, gamma 1-3, delta, and ep silon), we investigated the extent to which different CKI isoforms contribu te to NFAK activity. Using an NF-M-derived substrate, we determined that NF AK activity copurified with casein kinase activity through two purification steps. In an in-gel kinase assay, NFAK activity occurred at 36-40 kDa, cor responding to the size of CKI alpha isoforms. Chicken neurons express trans cripts encoding four alternatively spliced variants of CKI alpha (CKI alpha , CKI alpha S, CKI alpha L, and CKI alpha LS) differing in the presence or absence of two inserts, L and S. Using antibodies against different isoform s or with broad CKI specificity, we determined that all four CKI alpha vari ants, as well as other CKI family members, are present in chicken brain. Ho wever, only CKI alpha and CKI alpha S could be detected in purified NFAK. A lso, immunoprecipitation studies showed that CKI alpha and CKI alpha S toge ther account for NFAK activity. These findings raise the possibility that o nly a subset of CKI isoforms may be able to associate with and/or phosphory late NFs.