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STAT5A-deficient mice demonstrate a defect in granulocyte-macrophage colony-stimulating factor-induced proliferation
G M Feldman1, L A Rosenthal, X Liu
1Division of Cytokine Biology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892-4555, USA.
Abstract:
Responses of cells to cytokines typically involve the activation of a family of latent DNA binding proteins, referred to as signal transducers and activators of transcription (STAT) proteins, which are critical for the expression of early response genes. Of the seven known STAT proteins, STAT5 (originally called mammary gland factor) has been shown to be activated by several cytokines, such as granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5, which are known to play important roles in growth and differentiation of hematopoietic precursors. In this report we have used mice that are deficient in STAT5A (one of two homologues of STAT5) to study the role of STAT5A in GM-CSF stimulation of cells. When bone marrow-derived macrophages were generated by differentiation with macrophage-CSF (M-CSF), exposure of cells from wild-type mice to GM-CSF resulted in a typical pattern of assembly of DNA binding proteins specific for the gamma activation sequence (GAS) element within the beta-casein promoter. However, in cells from the STAT5A null mouse one of the shifted bands was absent. Immunoblotting analysis in the null mice showed that lack of STAT5A protein resulted in no alteration in activation of STAT5B by tyrosine phosphorylation. Proliferation experiments revealed that, when exposed to increasing concentrations of GM-CSF, cells derived from the null mice grew considerably more slowly than cells derived from the wild-type mice. Moreover, expression of GM-CSF-dependent genes, CIS and A1, was markedly inhibited in cells derived from null mice as compared with those of wild-type mice. The decreased expression observed with A1, a bcl-2 like gene, may account in part for the suppression of growth in cells from the null mice. These data suggest that the presence of STAT5A during the GM-CSF-induced assembly of STAT5 dimers is critical for the formation of competent transcription factors that are required for both gene expression and cell proliferation.
Insights
Signal transducers and activators of transcription 5A (STAT5A) are crucial for gene expression and cell proliferation. STAT5A deficiency impairs responses to granulocyte-macrophage colony-stimulating factor (GM-CSF), impacting hematopoietic precursor growth.
Area of Science:
- Cellular and Molecular Biology
- Immunology
- Hematopoiesis
Background:
- Signal transducers and activators of transcription (STAT) proteins mediate cellular responses to cytokines.
- STAT5, particularly STAT5A, is activated by cytokines like GM-CSF, IL-3, and IL-5, influencing hematopoietic precursor growth and differentiation.
Purpose of the Study:
- To investigate the specific role of STAT5A in GM-CSF-stimulated cellular responses using STAT5A-deficient mice.
- To elucidate the impact of STAT5A deficiency on gene expression and cell proliferation.
Main Methods:
- Generation of bone marrow-derived macrophages from STAT5A-deficient and wild-type mice.
- Analysis of DNA-binding protein assembly via gamma activation sequence (GAS) element in the beta-casein promoter.
- Immunoblotting to assess STAT5B activation.
- Cell proliferation assays and gene expression analysis of GM-CSF-dependent genes (CIS and A1).
Main Results:
- STAT5A deficiency resulted in the absence of a specific DNA-binding protein complex in response to GM-CSF.
- STAT5A deficiency did not affect STAT5B tyrosine phosphorylation.
- Cells from STAT5A-null mice exhibited significantly reduced proliferation and marked inhibition of CIS and A1 gene expression upon GM-CSF stimulation.
- Suppressed expression of the A1 gene, a bcl-2-like gene, may contribute to reduced cell growth.
Conclusions:
- STAT5A is essential for the formation of functional STAT5 dimers required for GM-CSF-induced gene expression and cell proliferation.
- STAT5A plays a critical role in mediating the biological effects of GM-CSF on hematopoietic cells.