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Inhibition of interleukin 3 and colony-stimulating factor 1-stimulated marrow cell proliferation by pertussis toxin
1Department of Internal Medicine, University of Virginia School of Medicine, Charlottesville 22908.
Abstract:
Pertussis toxin (PT) catalyzes the ADP-ribosylation of several guanine nucleotide-binding (G) proteins that are involved in the transduction of cell surface receptor-mediated signals. Involvement of such G-proteins in regulation of hematopoiesis by two growth factors, colony-stimulating factor-1 (CSF-1) and interleukin 3 (IL 3), was investigated using pertussis toxin. Continuous or pulse exposure of murine bone marrow cells to pertussis toxin inhibited CSF-1 or IL 3-induced colony formation by approximately 50%. Pertussis toxin inhibition was also demonstrated against partially separated marrow from 5-fluorouracil-treated mice. The toxin effect was blocked by heating (95 degrees C for 30 minutes), by antitoxin antibody and was not associated with increased cAMP levels in target cells. In experiments with murine marrow, toxin-mediated inhibition appeared to involve predominantly the macrophage lineage. IL 3 stimulation of proliferation of the murine marrow-derived factor-dependent cell line FDC-P1, as measured by 3H-TdR incorporation, and CSF-1 stimulation of pure populations of murine bone marrow derived macrophages, as measured by DNA content and cell number, was also inhibited. Analysis of the effects of pertussis toxin on the growth of single cells stimulated by IL 3 demonstrated that this inhibition involved a decreased growth rate rather than a toxic ablation of cells. Phorbol myristate acetate (PMA) stimulated FDC-P1 cells and was able to abrogate the PT inhibition of IL 3 stimulation of these cells, suggesting but not establishing that IL 3 may mediate its proliferative effects through activating protein kinase C.
Insights
Pertussis toxin (PT) inhibits hematopoietic growth factors like colony-stimulating factor-1 (CSF-1) and interleukin-3 (IL-3) by approximately 50%. This suggests G-protein involvement in regulating cell proliferation and differentiation.
Area of Science:
- Cell Biology
- Hematopoiesis
- Signal Transduction
Background:
- Pertussis toxin (PT) modifies guanine nucleotide-binding (G) proteins, crucial for cell signaling.
- Hematopoiesis, the formation of blood cellular components, is regulated by growth factors like CSF-1 and IL-3.
Purpose of the Study:
- To investigate the role of G-proteins in hematopoiesis regulated by CSF-1 and IL-3.
- To determine the effect of PT on growth factor-induced hematopoietic cell proliferation and differentiation.
Main Methods:
- Murine bone marrow cells and specific cell lines (FDC-P1) were exposed to PT.
- Colony formation assays and cell proliferation measurements (3H-TdR incorporation, DNA content) were performed.
- PT inhibition was assessed by heat inactivation, antitoxin antibodies, and cAMP level analysis.
Main Results:
- PT inhibited CSF-1 and IL-3-induced colony formation by ~50%.
- PT reduced proliferation in FDC-P1 cells and CSF-1 stimulated macrophages, primarily affecting the macrophage lineage.
- Inhibition was due to decreased growth rate, not cell death, and could be partially reversed by PMA.
Conclusions:
- G-proteins are involved in CSF-1 and IL-3-mediated hematopoiesis.
- PT significantly inhibits hematopoietic cell proliferation, particularly in the macrophage lineage.
- IL-3 may utilize protein kinase C pathways for proliferation, modulated by G-proteins.