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The role of protein kinase C isotypes in the proliferation and differentiation of human leukemia cells

Murray, Nicole Renee
http://rave.ohiolink.edu/etdc/view?acc_num=case1058212510

Abstract Details

1995, Doctor of Philosophy, Case Western Reserve University, Pharmacology.
Protein kinase C (PKC) is a family of serine/threonine kinases known to be involved in the proliferation and differentiation of human leukemia cells. The present study is an investigation of the role of individual PKC isotypes in leukemic proliferation and PMA-induced differentiation and the molecular basis for PKC isotype-specific function. Proliferating human erythroleukemia (K562) cells express the α, βII and ζ PKC isotypes. The PKC activator, phorbol myristate acetate (PMA) induces megakaryocytic differentiation and cytostasis in these cells resulting in a decrease in βII PKC levels and increases in α and ζ PKC levels. K562 cells overexpressing a PKC grow more slowly and are more sensitive to the cytostatic effects of PMA than control cells whereas cells overexpressing βII PKC are less sensitive to PMA. Removal of PMA and resumption of proliferation correlates with re-expression of βII PKC to near control levels. Proliferation of PMA-withdrawn cells can be partially inhibited by anti-sense βII PKC oligodeoxyribonucleotides which significantly inhibit re-expression of βII PKC. These data demonstrate that βII PKC is req uired for K562 cell proliferation whereas α PKC is involved in cytostasis and/or megakaryocytic differentiation. The cytostatic effects of PMA can be overcome by co-administration of another PKC activator, bryostatin 1 (bryo) which promotes continued cellular proliferation. The proliferative effect of bryo correlates with selective translocation of βII PKC to the nucleus, where it directly phosphorylates lamin B at two consensus PKC phosphorylation sites. The molecular basis for βII PKC-selective nuclear activation and lamin B phosphorylation was investigated using an in-vitro reconstitution system. βII PKC phosphorylates nuclear envelope lamin B at 10-20 times the rate of α PKC whereas both kinases phosphorylate soluble lamin B at similar rates on the same peptides. 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 polymerized lamin B. Nuclear membrane extracts reconstitute βII PKC-selective phosphorylation of extracted nuclear lamina and selectively activate βII PKC histone kinase activity, indicating the presence of a βII PKC-selective nuclear membrane activation factor (NMAF). NMAF activity is not affected by exhaustive protease treatment, suggesting that it is a nuclear membrane lipid(s) or lipid metabolite. To investigate the molecular basis for α PKC function in cellular differentiation and cytostasis, we constructed two chimera of α and βII PKC in which the regulatory and catalytic domains are exchanged. Expression of the βII/α (regulatory/catalytic) PKC chimer in K562 cells induces cytostasis and increases sensitivity to megakaryocytic differentiation as seen in cells overexpressing α PKC. In contrast, expression of the α /βII PKC chimer has no effect on these parameters. These results indicate that the molecular determinants necessary for α PKC-mediated cytostasis reside in the catalytic domain and suggest that this domain is a potential therapeutic target in the treatment of human leukemi
Alan Fields (Advisor)
123 p.
protein kinase C isotypes; human leukemia cells

Recommended Citations

Citations

  • Murray, N. R. (1995). The role of protein kinase C isotypes in the proliferation and differentiation of human leukemia cells [Doctoral dissertation, Case Western Reserve University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=case1058212510

    APA Style (7th edition)

  • Murray, Nicole. The role of protein kinase C isotypes in the proliferation and differentiation of human leukemia cells. 1995. Case Western Reserve University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=case1058212510.

    MLA Style (8th edition)

  • Murray, Nicole. "The role of protein kinase C isotypes in the proliferation and differentiation of human leukemia cells." Doctoral dissertation, Case Western Reserve University, 1995. http://rave.ohiolink.edu/etdc/view?acc_num=case1058212510

    Chicago Manual of Style (17th edition)

case1058212510
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© 1995, all rights reserved.
This open access ETD is published by Case Western Reserve University School of Graduate Studies and OhioLINK.