A Cell Type-specific Constitutive Point Mutant of the Common β-Subunit of the Human Granulocyte-Macrophage Colony-stimulating Factor (GM-CSF), Interleukin (IL)-3, and IL-5 Receptors Requires the GM-CSF Receptor α-Subunit for Activation

Brendan J Jenkins,Fei Le,Thomas J Gonda

doi:10.1074/jbc.274.13.8669

Brendan J Jenkins, Fei Le + Show 1 more

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https://doi.org/10.1074/jbc.274.13.8669

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Abstract

The high affinity receptor for human granulocyte-macrophage colony-stimulating factor (GM-CSF) consists of a cytokine-specific alpha-subunit (hGMRalpha) and a common signal-transducing beta-subunit (hbetac) that is shared with the interleukin-3 and -5 receptors. We have previously identified a constitutively active extracellular point mutant of hbetac, I374N, that can confer factor independence on murine FDC-P1 cells but not BAF-B03 or CTLL-2 cells (Jenkins, B. J., D'Andrea, R. J., and Gonda, T. J. (1995) EMBO J. 14, 4276-4287). This restricted activity suggested the involvement of cell type-specific signaling molecules in the activation of this mutant. We report here that one such molecule is the mouse GMRalpha (mGMRalpha) subunit, since introduction of mGMRalpha, but not hGMRalpha, into BAF-B03 or CTLL-2 cells expressing the I374N mutant conferred factor independence. Experiments utilizing mouse/human chimeric GMRalpha subunits indicated that the species specificity lies in the extracellular domain of GMRalpha. Importantly, the requirement for mGMRalpha correlated with the ability of I374N (but not wild-type hbetac) to constitutively associate with mGMRalpha. Expression of I374N in human factor-dependent UT7 cells also led to factor-independent proliferation, with concomitant up-regulation of hGMRalpha surface expression. Taken together, these findings suggest a critical role for association with GMRalpha in the constitutive activity of I374N.

Highlights

GM-CSF1 is a potent cytokine that promotes the survival, proliferation, differentiation, and functional activity of a wide variety of hemopoietic cell types including monocytes/macrophages, granulocytes, and myeloid progenitor cells
This mutant was unable to confer factor independence on mouse IL-3-dependent BAF-B03 cells, leading us to suggest that the cell type-specific activity of this mutant may reflect the presence of a ␤-subunit-associated signaling molecule in FDC-P1 cells, but not in BAF-B03 cells, that is required by this mutant for constitutive activation [16]
Constitutive Activation of I374N in Mouse Cells Requires mGMR␣—The I374N mutation in the extracellular domain of h␤c confers factor independence on mouse FDC-P1 cells but not BAF-B03 or CTLL-2 cells [16], raising the possibility that cell type-specific signaling molecules are involved in its activation

Summary

Introduction

GM-CSF1 is a potent cytokine that promotes the survival, proliferation, differentiation, and functional activity of a wide variety of hemopoietic cell types including monocytes/macrophages, granulocytes, and myeloid progenitor cells (reviewed in Ref. 1). Isolation of Factor-independent BAF/I374N Cells Infected with an FDC-P1 cDNA Retroviral Expression Library—We have previously identified a constitutively activating point mutation, I374N, in the extracellular region of h␤c by virtue of its ability to confer factor-independent growth on FDC-P1 cells [16].

Results

Conclusion