Activation or inactivation mechanism of brain cdc2-like kinase, cdk5

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Cyclin-dependent kinases are key factors in progression of cell cycle in eukaryotic cells. CDK5 is an unique CDK whosc kinase activity is detected only in post-mitotic neurons. Several lines of evidence suggest that CDK5 involves in neuronal migration or positioning during development via phosphorylation of axonal cytoskeletons such as microtubule-associated protein tau and neurofilaments. However, it is not known how its kinase activity is regulated. We investigated here the activation or inactivation mechanism of CDK5.1. We studied the activation mechanism of CDK5 in primary cultured neurons using the phosphorylation of neurofilament-H subunit as a marker. CDK5 was activated at 4 to 5 days after plating when phosphorylated form of neurofilament-H subunit appeared and synapse formation occurred. Addition of BDNF into culture medium induced activation of CDK5. These results suggest that activation of CDK5 was induced by increased secretion of BDNF after synapse formation.2. CDK5 is activated by binding to activating subunit p35 which is expressed in neurons exclusively. p35 is a short life protein degraded in a few hours. CDK5 decreased its activity along with degradation of p35. p35 was degraded by proteasome as well as cyclins in proliferating cells. Phosphorylation was suggested to be a signal for degradation.3. p35 is proteolysed to p25 during purification of CDK5 from bovine or porcine brains. Proteases involving in this limited proteolysis was shown to be calpain. Addition of CaィイD12+ィエD1 to porcine brain extract induced proteolysis of p35 to p25. This proteolysis was suppressed by calpain inhibitor. The proteolysis changed the solubility of CDK5. P35/CDK5 precipitated by brief centrifugation became to be soluble after proteolytic conversion to p25. Limited proteolysis of p35 to p25 was enhanced at the time of neuronal cell death.