Abstract
To determine if MYB protein is preferentially required during specific stages of normal human hematopoiesis we incubated normal marrow mononuclear cells (MNC) with c-myb antisense oligodeoxynucleotides. Treated cells were cultured in semisolid medium under conditions designed to favor the growth of specific progenitor cell types. Compared with untreated controls, granulocyte-macrophage (GM) CFU-derived colonies decreased 77% when driven by recombinant human (rH) IL-3, and 85% when stimulated by rH GM colony-stimulating factor (CSF); erythroid burst-forming unit (BFU-E)- and CFU-E-derived colonies decreased 48 and 78%, respectively. In contrast, numbers of G-CSF-stimulated granulocyte colonies derived from antisense treated MNC were unchanged from controls, though the numbers of cells composing these colonies decreased approximately 90%. Similar results were obtained when MY10+ cells were exposed to c-myb antisense oligomers. When compared with untreated controls, numbers of CFU-GM and BFU-E colonies derived from MY10+ cells were unchanged, but the numbers of cells composing these colonies were reduced approximately 75 and greater than 90%, respectively, in comparison with controls. c-myc sense and antisense oligomers were without significant effect in these assays. Using the reverse transcription-polymerase chain reaction, c-myb mRNA was detected in developing hematopoietic cells on days 0-8. At day 14 c-myb expression was no longer detectable using this technique. These results suggest that c-myb is required for proliferation of intermediate-late myeloid and erythroid progenitors, but is less important for lineage commitment and early progenitor cell amplification.
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Selected References
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- Anfossi G., Gewirtz A. M., Calabretta B. An oligomer complementary to c-myb-encoded mRNA inhibits proliferation of human myeloid leukemia cell lines. Proc Natl Acad Sci U S A. 1989 May;86(9):3379–3383. doi: 10.1073/pnas.86.9.3379. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bradley T. R., Metcalf D. The growth of mouse bone marrow cells in vitro. Aust J Exp Biol Med Sci. 1966 Jun;44(3):287–299. doi: 10.1038/icb.1966.28. [DOI] [PubMed] [Google Scholar]
- Brandt J., Baird N., Lu L., Srour E., Hoffman R. Characterization of a human hematopoietic progenitor cell capable of forming blast cell containing colonies in vitro. J Clin Invest. 1988 Sep;82(3):1017–1027. doi: 10.1172/JCI113658. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Caracciolo D., Shirsat N., Wong G. G., Lange B., Clark S., Rovera G. Recombinant human macrophage colony-stimulating factor (M-CSF) requires subliminal concentrations of granulocyte/macrophage (GM)-CSF for optimal stimulation of human macrophage colony formation in vitro. J Exp Med. 1987 Dec 1;166(6):1851–1860. doi: 10.1084/jem.166.6.1851. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Caracciolo D., Valtieri M., Venturelli D., Peschle C., Gewirtz A. M., Calabretta B. Lineage-specific requirement of c-abl function in normal hematopoiesis. Science. 1989 Sep 8;245(4922):1107–1110. doi: 10.1126/science.2672339. [DOI] [PubMed] [Google Scholar]
- Civin C. I., Strauss L. C., Brovall C., Fackler M. J., Schwartz J. F., Shaper J. H. Antigenic analysis of hematopoiesis. III. A hematopoietic progenitor cell surface antigen defined by a monoclonal antibody raised against KG-1a cells. J Immunol. 1984 Jul;133(1):157–165. [PubMed] [Google Scholar]
- Clark S. C., Kamen R. The human hematopoietic colony-stimulating factors. Science. 1987 Jun 5;236(4806):1229–1237. doi: 10.1126/science.3296190. [DOI] [PubMed] [Google Scholar]
- Clarke M. F., Kukowska-Latallo J. F., Westin E., Smith M., Prochownik E. V. Constitutive expression of a c-myb cDNA blocks Friend murine erythroleukemia cell differentiation. Mol Cell Biol. 1988 Feb;8(2):884–892. doi: 10.1128/mcb.8.2.884. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dash P., Lotan I., Knapp M., Kandel E. R., Goelet P. Selective elimination of mRNAs in vivo: complementary oligodeoxynucleotides promote RNA degradation by an RNase H-like activity. Proc Natl Acad Sci U S A. 1987 Nov;84(22):7896–7900. doi: 10.1073/pnas.84.22.7896. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ferrero D., Broxmeyer H. E., Pagliardi G. L., Venuta S., Lange B., Pessano S., Rovera G. Antigenically distinct subpopulations of myeloid progenitor cells (CFU-GM) in human peripheral blood and marrow. Proc Natl Acad Sci U S A. 1983 Jul;80(13):4114–4118. doi: 10.1073/pnas.80.13.4114. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gewirtz A. M., Anfossi G., Venturelli D., Valpreda S., Sims R., Calabretta B. G1/S transition in normal human T-lymphocytes requires the nuclear protein encoded by c-myb. Science. 1989 Jul 14;245(4914):180–183. doi: 10.1126/science.2665077. [DOI] [PubMed] [Google Scholar]
- Gewirtz A. M., Calabretta B. A c-myb antisense oligodeoxynucleotide inhibits normal human hematopoiesis in vitro. Science. 1988 Dec 2;242(4883):1303–1306. doi: 10.1126/science.2461588. [DOI] [PubMed] [Google Scholar]
- Gonda T. J., Metcalf D. Expression of myb, myc and fos proto-oncogenes during the differentiation of a murine myeloid leukaemia. Nature. 1984 Jul 19;310(5974):249–251. doi: 10.1038/310249a0. [DOI] [PubMed] [Google Scholar]
- Heikkila R., Schwab G., Wickstrom E., Loke S. L., Pluznik D. H., Watt R., Neckers L. M. A c-myc antisense oligodeoxynucleotide inhibits entry into S phase but not progress from G0 to G1. 1987 Jul 30-Aug 5Nature. 328(6129):445–449. doi: 10.1038/328445a0. [DOI] [PubMed] [Google Scholar]
- Holt J. T., Redner R. L., Nienhuis A. W. An oligomer complementary to c-myc mRNA inhibits proliferation of HL-60 promyelocytic cells and induces differentiation. Mol Cell Biol. 1988 Feb;8(2):963–973. doi: 10.1128/mcb.8.2.963. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ichikawa Y., Pluznik D. H., Sachs L. In vitro control of the development of macrophage and granulocyte colonies. Proc Natl Acad Sci U S A. 1966 Aug;56(2):488–495. doi: 10.1073/pnas.56.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Iscove N. N. The role of erythropoietin in regulation of population size and cell cycling of early and late erythroid precursors in mouse bone marrow. Cell Tissue Kinet. 1977 Jul;10(4):323–334. doi: 10.1111/j.1365-2184.1977.tb00300.x. [DOI] [PubMed] [Google Scholar]
- Kastan M. B., Slamon D. J., Civin C. I. Expression of protooncogene c-myb in normal human hematopoietic cells. Blood. 1989 May 1;73(6):1444–1451. [PubMed] [Google Scholar]
- Lipton J. M., Kudisch M., Nathan D. G. Response of three classes of human erythroid progenitors to the absence of erythropoietin in vitro as a measure of progenitor maturity. Exp Hematol. 1981 Nov;9(10):1035–1041. [PubMed] [Google Scholar]
- Majello B., Kenyon L. C., Dalla-Favera R. Human c-myb protooncogene: nucleotide sequence of cDNA and organization of the genomic locus. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9636–9640. doi: 10.1073/pnas.83.24.9636. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Metcalf D. The granulocyte-macrophage colony-stimulating factors. Science. 1985 Jul 5;229(4708):16–22. doi: 10.1126/science.2990035. [DOI] [PubMed] [Google Scholar]
- Prochownik E. V., Kukowska J., Rodgers C. c-myc antisense transcripts accelerate differentiation and inhibit G1 progression in murine erythroleukemia cells. Mol Cell Biol. 1988 Sep;8(9):3683–3695. doi: 10.1128/mcb.8.9.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rappolee D. A., Mark D., Banda M. J., Werb Z. Wound macrophages express TGF-alpha and other growth factors in vivo: analysis by mRNA phenotyping. Science. 1988 Aug 5;241(4866):708–712. doi: 10.1126/science.3041594. [DOI] [PubMed] [Google Scholar]
- Sachs L. The molecular control of blood cell development. Science. 1987 Dec 4;238(4832):1374–1379. doi: 10.1126/science.3317831. [DOI] [PubMed] [Google Scholar]
- Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. doi: 10.1126/science.2448875. [DOI] [PubMed] [Google Scholar]
- Sieff C. A. Hematopoietic growth factors. J Clin Invest. 1987 Jun;79(6):1549–1557. doi: 10.1172/JCI112988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tebbi K., Rubin S., Cowan D. H., McCulloch E. A. A comparison of granulopoiesis in culture from blood and marrow cells of nonleukemic individuals and patients with acute leukemia. Blood. 1976 Aug;48(2):235–243. [PubMed] [Google Scholar]
- Till J. E., McCulloch E. A. Hemopoietic stem cell differentiation. Biochim Biophys Acta. 1980 Nov 26;605(4):431–459. doi: 10.1016/0304-419x(80)90009-8. [DOI] [PubMed] [Google Scholar]
- Watt R., Stanton L. W., Marcu K. B., Gallo R. C., Croce C. M., Rovera G. Nucleotide sequence of cloned cDNA of human c-myc oncogene. Nature. 1983 Jun 23;303(5919):725–728. doi: 10.1038/303725a0. [DOI] [PubMed] [Google Scholar]
- Weinberg R. A. The action of oncogenes in the cytoplasm and nucleus. Science. 1985 Nov 15;230(4727):770–776. doi: 10.1126/science.2997917. [DOI] [PubMed] [Google Scholar]
- Westin E. H., Gallo R. C., Arya S. K., Eva A., Souza L. M., Baluda M. A., Aaronson S. A., Wong-Staal F. Differential expression of the amv gene in human hematopoietic cells. Proc Natl Acad Sci U S A. 1982 Apr;79(7):2194–2198. doi: 10.1073/pnas.79.7.2194. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zamecnik P. C., Goodchild J., Taguchi Y., Sarin P. S. Inhibition of replication and expression of human T-cell lymphotropic virus type III in cultured cells by exogenous synthetic oligonucleotides complementary to viral RNA. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4143–4146. doi: 10.1073/pnas.83.12.4143. [DOI] [PMC free article] [PubMed] [Google Scholar]