Presence of a βII Protein Kinase C-selective Nuclear Membrane Activation Factor in Human Leukemia Cells

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In human promyelocytic (HL60) leukemia cells βII protein kinase C (PKC) is selectively translocated to the nucleus in response to proliferative stimuli. At the nucleus, βII PKC directly phosphorylates the nuclear envelope polypeptide lamin B at two consensus PKC phosphorylation sites, Ser395 and Ser405. Phosphorylation of these sites by βII PKC leads to solubilization of lamin B indicative of mitotic nuclear envelope breakdown in vitro (Hocevar, B. A., Burns, D. J., and Fields, A. P. (1993) J. Biol. Chem. 268, 7545-7552). We have now investigated the molecular basis for βII PKC-selective nuclear translocation and lamin B phosphorylation using an in vitro reconstitution system. We find that βII PKC phosphorylates nuclear envelope lamin B at 10-20 times the rate of a PKC, whereas both kinases phosphorylate soluble lamin B at similar rates. Comparative tryptic phosphopeptide analysis demonstrates that a PKC and βII PKC phosphorylate identical sites, Ser395 and Ser405, on soluble lamin B. These data suggest that a component(s) of the nuclear envelope confers βII PKC-selective nuclear activation and lamin B phosphorylation. Extraction of nuclear envelopes with either non-ionic detergent (2% n-octyl glucoside) or organic solvent (CHCl3/ CH3OH/H2O; 10:10:3) abolishes βII PKC-selective phosphorylation of nuclear lamin B. Nuclear membrane extracts reconstitute βII PKC-selective phosphorylation, indicating the presence of a βII PKC-selective nuclear membrane activation factor (NMAF). NMAF selectively activates βII PKC histone H1 kinase activity 3-4-fold above the level achieved with optimal concentrations of Ca2+, diacylglycerol, and phosphatidylserine. Finally, NMAF activity is not affected by exhaustive protease treatment, suggesting that it is a nuclear membrane lipid(s) or lipid metabolite. These data suggest that NMAF plays a physiologic role in the nuclear activation of βII PKC.

Original languageEnglish (US)
Pages (from-to)21385-21390
Number of pages6
JournalJournal of Biological Chemistry
Issue number33
StatePublished - Aug 19 1994

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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