Polyamine-activated protein kinase reaction from nuclei and nucleoli of Physarum polycephalum which phosphorylates a unique Mr 70 000 nonhistone protein.

Methods are described for the detection and purification of a protein kinase from nuclei and nucleoli of Physarum polycephalum which catalyzed transfer of phosphate from [gamma-32P]ATP to a unique nonhistone protein of Mr 70 000 in a reaction that was polyamine dependent. Enzymatic phosphorylation of the nonhistone protein by the purified protein kinase was stimulated greatly, at times more than 60-fold, by the polyamines spermidine and spermine. This unique polyamine-dependent reaction was localized on the rDNA minichromosome of the nucleolus. The polyamine-dependent protein kinase, which was first partially purified with the acidic nonhistone protein fraction from isolated nucleoli, was resolved from at least six other protein kinases by phosphocellulose chromatography into a catalytic component of Mr 26 000 and a complex comprised of the catalytic component associated with a phosphate acceptor protein of Mr 70 000. The complex also catalyzed polyamine-dependent phosphorylation of the endogenous Mr 70 000 component. The resolved catalytic component catalyzed polyamine-dependent phosphorylation of a dephosphorylated Mr 70 000 nonhistone protein that had been independently isolated from nucleoli and previously demonstrated to have properties concordant with a specific regulatory role in rRNA gene transcription [Keuhn, G. D., Affolter, H. U., Atmar, V. J., Seebeck, T., Gubler, U., & Braun, R. (1979) Proc. Natl. Acad. Sci. U.S.A. 76, 2541-2545]. These studies indicate one way that the polyamines may regulate rRNA gene transcription through the mediation of a highly specific nonhistone protein kinase.