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

Abstract

Methods are described for the detection and purification of a protein kinase from nuclei and nucleoli of P. polycephalum which catalyzed transfer of phosphate from [.gamma.-32P]ATP to a unique nonhistone protein of MW 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 6 other protein kinases by phosphocellulose chromatography into a catalytic component of MW 26,000 and a complex comprised of the catalytic component associated with a phosphate acceptor protein of MW 70,000. The complex catalyzed polyamine-dependent phosphorylation of the endogenous MW 70,000 component. The resolved catalytic component catalyzed polyamine-dependent phosphorylation of a dephosphorylated MW 70,000 nonhistone protein that was independently isolated from nucleoli and previously demonstrated to have properties concordant with a specific regulatory eole in rRNA gene transcription [Kuehn, G.D. et al.]. The polyamines may regulate rRNA gene transcription through the mediation of a highly specific nonhistone protein kinase.