Abstract
Initial studies revealed that proline-directed protein kinase FA(PDPK FA) was overexpressed in various cancerous tissues relative to normal controls. However, the functional role of overexpressed PDPK FAin cancer remains to be established. In this report, we explore the potential role of PDPK FAin leukaemia cell growth by investigating the effects of partial inhibition of this kinase on the malignant phenotype of human chronic myeloid leukaemia cells (K562). Cloning of PDPK FAcDNA and its recombinant antisense expression vector and PDPK FA-specific antibody were successfully developed. Two stable antisense clones of K562 cells were subcloned which expressed 70% and 45% of PDPK FArespectively, compared with control-transfected clone in both immunoprecipitate activity assay and immunoblot analysis. In sharp contrast, these two antisense clones expressed no significant suppression of any other related PDPK family members, indicating the specificity of these two antisense clones. Moreover, these antisense clones proportionally and potentially exhibited cell growth retardation, poor clonogenic growth in soft agar and loss of serum independence. The results demonstrate that specific antisense suppression of PDPK FAis sufficient to interfere with the growth of K562 cells, indicating that PDPK FAis essential for human chronic myeloid leukaemia cell growth. © 2000 Cancer Research Campaign
Keywords: chronic myeloid leukaemia cells K562, PDPK FA, cell growth
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- Bagchi M., Van Wijnen A., Katar M., Merriman H., Lian J., Stein J., Stein G., Maisel H. Sequence-specific DNA binding activities of nuclear matrix proteins of mammalian lens epithelial cells. J Cell Biochem. 1995 May;58(1):1–5. doi: 10.1002/jcb.240580102. [DOI] [PubMed] [Google Scholar]
- Freedman V. H., Shin S. I. Cellular tumorigenicity in nude mice: correlation with cell growth in semi-solid medium. Cell. 1974 Dec;3(4):355–359. doi: 10.1016/0092-8674(74)90050-6. [DOI] [PubMed] [Google Scholar]
- Gillespie P. G., Hudspeth A. J. Chemiluminescence detection of proteins from single cells. Proc Natl Acad Sci U S A. 1991 Mar 15;88(6):2563–2567. doi: 10.1073/pnas.88.6.2563. [DOI] [PMC free article] [PubMed] [Google Scholar]
- He X., Saint-Jeannet J. P., Woodgett J. R., Varmus H. E., Dawid I. B. Glycogen synthase kinase-3 and dorsoventral patterning in Xenopus embryos. Nature. 1995 Apr 13;374(6523):617–622. doi: 10.1038/374617a0. [DOI] [PubMed] [Google Scholar]
- Hemmings B. A., Yellowlees D., Kernohan J. C., Cohen P. Purification of glycogen synthase kinase 3 from rabbit skeletal muscle. Copurification with the activating factor (FA) of the (Mg-ATP) dependent protein phosphatase. Eur J Biochem. 1981 Oct;119(3):443–451. doi: 10.1111/j.1432-1033.1981.tb05628.x. [DOI] [PubMed] [Google Scholar]
- Holley R. W., Kiernan J. A. "Contact inhibition" of cell division in 3T3 cells. Proc Natl Acad Sci U S A. 1968 May;60(1):300–304. doi: 10.1073/pnas.60.1.300. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jeha S., Luo X. N., Beran M., Kantarjian H., Atweh G. F. Antisense RNA inhibition of phosphoprotein p18 expression abrogates the transformed phenotype of leukemic cells. Cancer Res. 1996 Mar 15;56(6):1445–1450. [PubMed] [Google Scholar]
- Lee S. C., Kuan C. Y., Yang C. C., Yang S. D. Bioflavonoids commonly and potently induce tyrosine dephosphorylation/inactivation of oncogenic proline-directed protein kinase FA in human prostate carcinoma cells. Anticancer Res. 1998 Mar-Apr;18(2A):1117–1121. [PubMed] [Google Scholar]
- Lozzio C. B., Lozzio B. B. Human chronic myelogenous leukemia cell-line with positive Philadelphia chromosome. Blood. 1975 Mar;45(3):321–334. [PubMed] [Google Scholar]
- Plyte S. E., Hughes K., Nikolakaki E., Pulverer B. J., Woodgett J. R. Glycogen synthase kinase-3: functions in oncogenesis and development. Biochim Biophys Acta. 1992 Dec 16;1114(2-3):147–162. doi: 10.1016/0304-419x(92)90012-n. [DOI] [PubMed] [Google Scholar]
- Reichlin M. Use of glutaraldehyde as a coupling agent for proteins and peptides. Methods Enzymol. 1980;70(A):159–165. doi: 10.1016/s0076-6879(80)70047-2. [DOI] [PubMed] [Google Scholar]
- Shin S. I., Freedman V. H., Risser R., Pollack R. Tumorigenicity of virus-transformed cells in nude mice is correlated specifically with anchorage independent growth in vitro. Proc Natl Acad Sci U S A. 1975 Nov;72(11):4435–4439. doi: 10.1073/pnas.72.11.4435. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Todaro G. J., De Larco J. E. Growth factors produced by sarcoma virus-transformed cells. Cancer Res. 1978 Nov;38(11 Pt 2):4147–4154. [PubMed] [Google Scholar]
- Vandenheede J. R., Yang S. D., Goris J., Merlevede W. ATP x Mg-dependent protein phosphatase from rabbit skeletal muscle. II. Purification of the activating factor and its characterization as a bifunctional protein also displaying synthase kinase activity. J Biol Chem. 1980 Dec 25;255(24):11768–11774. [PubMed] [Google Scholar]
- Woodgett J. R. Molecular cloning and expression of glycogen synthase kinase-3/factor A. EMBO J. 1990 Aug;9(8):2431–2438. doi: 10.1002/j.1460-2075.1990.tb07419.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Yang C. C., Hsu C. P., Sheu J. C., Mai X. Y., Yang S. D. Differential tyrosine phosphorylation/activation of oncogenic proline-directed protein kinase F(A)/GSK-3alpha in well and poorly differentiated human prostate carcinoma cells. J Protein Chem. 1998 May;17(4):329–335. doi: 10.1023/a:1022503215525. [DOI] [PubMed] [Google Scholar]
- Yang S. D., Yu J. S., Lee T. T., Ni M. H., Yang C. C., Ho Y. S., Tsen T. Z. Association of protein kinase FA/GSK-3alpha (a proline-directed kinase and a regulator of protooncogenes) with human cervical carcinoma dedifferentiation/progression. J Cell Biochem. 1995 Oct;59(2):143–150. doi: 10.1002/jcb.240590203. [DOI] [PubMed] [Google Scholar]
- Yang S. D., Yu J. S., Wen Z. D. Tumor promoter phorbol ester reversibly modulates tyrosine dephosphorylation/inactivation of protein kinase FA/GSK-3 alpha in A431 cells. J Cell Biochem. 1994 Dec;56(4):550–558. doi: 10.1002/jcb.240560416. [DOI] [PubMed] [Google Scholar]
- Yang S. D., Yu J. S., Yang C. C., Lee S. C., Lee T. T., Ni M. H., Kuan C. Y., Chen H. C. Overexpression of protein kinase FA/GSK-3 alpha (a proline-directed protein kinase) correlates with human hepatoma dedifferentiation/progression. J Cell Biochem. 1996 May;61(2):238–245. doi: 10.1002/(SICI)1097-4644(19960501)61:2%3C238::AID-JCB7%3E3.0.CO;2-V. [DOI] [PubMed] [Google Scholar]
- Yu J. S., Yang S. D. Tyrosine dephosphorylation and concurrent inactivation of protein kinase FA/GSK-3 alpha by genistein in A431 cells. J Cell Biochem. 1994 Sep;56(1):131–141. doi: 10.1002/jcb.240560117. [DOI] [PubMed] [Google Scholar]