Abstract Incubation of isolated, Triton X-100 washed rat liver nuclei with purified bovine myocardial calpain II resulted in solubilization of a histone H1 kinase activity. The release of kinase from nuclei could be prevented by including the calpain inhibitors leupeptin or calpastatin in the incubation. Titration with Ca 2+/EGTA buffers indicated that the calpain-dependent release of the kinase was half-maximal at approximately 3 μM Ca 2+. In contrast, calpain II required at least 50 μM Ca 2+ to produce detectable proteolysis of soluble or membrane-associated substrates. These results suggest that the cell nucleus is a site of calpain II activation and function.