Cloning of three novel neuronal Cdk5 activator binding proteins

Neuronal Cdc2-like kinase (Nclk) is involved in the regulation of neuronal differentiation and neuro-cytoskeleton dynamics. The active kinase consists of a catalytic subunit, Cdk5, and a 25 kDa activator protein (p25(nck5a)) derived from a 35 kDa neuronal-specific protein (p35(nck5a)). As an extensi on of our previous study (Qi, Z., Tang, D., Zhu, X., Fujita, D.J., Wang, J. H., 1998. Association of neurofilament proteins with neuronal Cdk5 activato r. J. Biol. Chem. 270, 2329-2335), which showed that neurofilament is one o f the p35(nck5a)-associated proteins, we now report the isolation of three other novel p35(nck5a)-associated proteins using the yeast two-hybrid scree n. The full-length forms of these three novel proteins, designated C42, C48 and C53, have a molecular mass of 66, 24, and 57 kDa, respectively. Northe rn analysis indicates that these novel proteins are widely expressed in hum an tissues, including the heart, brain, skeletal muscle, placenta? lung, li ver, kidney and pancreas. The bacterially expressed glutathione S-transfera se (GST)-fusion forms of these three proteins were able to co-precipitate p 35(nck5a) complexed with Cdk5 from insect cell lysate. Among these three pr oteins, only C48 and C53 can be phosphorylated by Nclk, suggesting that the y may be the substrates of Nclk. Sequence homology searches have suggested that the C48 protein is marginally related to restin protein, whereas the C 42 protein has homologues of unknown function in Caenorhabditis elegans and Arabidopsis thaliana. (C) 2000 Elsevier Science B.V. All rights reserved.