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 has properties consistent with casein kinase I (CKI). Because CKI consists of a family of as many as seven genes (alpha, beta, gamma1-3, delta, and epsilon), we investigated the extent to which different CKI isoforms contribute to NFAK activity. Using an NF-M-derived substrate, we determined that NFAK activity copurified with casein kinase activity through two purification steps. In an in-gel kinase assay, NFAK activity occurred at 36-40 kDa, corresponding to the size of CKIalpha isoforms. Chicken neurons express transcripts encoding four alternatively spliced variants of CKIalpha (CKIalpha, CKIalphaS, CKIalphaL, and CKIalphaLS) differing in the presence or absence of two inserts, L and S. Using antibodies against different isoforms or with broad CKI specificity, we determined that all four CKIalpha variants, as well as other CKI family members, are present in chicken brain. However, only CKIalpha and CKIalphaS could be detected in purified NFAK. Also, immunoprecipitation studies showed that CKIalpha and CKIalphaS together account for NFAK activity. These findings raise the possibility that only a subset of CKI isoforms may be able to associate with and/or phosphorylate NFs.
Fu
Z
Z
Department of Anatomy and Cell Biology, University of Florida College of Medicine, Gainesville 32610-0235, USA.
Green
C L
CL
Bennett
G S
GS
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology*; D047388:Casein Kinases; D002645:Chickens; D004789:Enzyme Activation; D015151:Immunoblotting; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016900:Neurofilament Proteins / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D007527:Isoenzymes; D016900:Neurofilament Proteins; C061195:neurofilament protein M; D011494:Protein Kinases; C084357:neurofilament-associated kinase; D047388:Casein Kinases