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Biological characterization of uncleavable plasma membrane-anchored human macrophage colony-stimulating factor
P Deng1, Y L Wang, V L Shahbazian
1Department of Pathology, Childrens Hospital of Los Angeles, Los Angeles, California 90027, USA.
Abstract:
The cell-surface form of human macrophage colony-stimulating factor (CSF-1(256), M-CSFalpha) is a plasma membrane-anchored transmembrane protein from which the soluble CSF-1 is released by ectodomain proteolytic cleavage. We have previously generated two forms of cell surface CSF-1 which failed to undergo the cleavage by deleting residues 161-165 or residues 159-165 in the extracellular juxtamembrane region (1). To determine the biologic significance of the ectodomain cleavage, we compared the biosynthesis and biologic activities of uncleavable mutant CSF-1 forms with those of the cleavable wild-type (WT) CSF-1. We found that the uncleavable CSF-1 forms were able to accumulate on cell surface at about threefold higher level than the cleavable WT CSF-1 did. We further demonstrated that the uncleavable plasma membrane-anchored forms of CSF-1 were biologically active in mediating the proliferation of CSF-1-dependent cells as well as the intercellular adhesion between CSF-1 receptor-bearing cells and CSF-1 expressing cells. Furthermore, the adhesive activity of uncleavable CSF-1 forms was about twofold stronger than that of WT CSF-1, which indicated that the ectodomain cleavage system plays an important role in regulating the biologic activities of membrane-anchored CSF-1.
Insights
Cell surface human macrophage colony-stimulating factor (CSF-1) ectodomain cleavage regulates its biological activity. Preventing cleavage increases CSF-1 cell surface accumulation and enhances cell proliferation and adhesion.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Cell-surface human macrophage colony-stimulating factor (CSF-1) is a transmembrane protein.
- Soluble CSF-1 is released via ectodomain proteolytic cleavage.
- Previous studies generated uncleavable CSF-1 mutants by deleting specific extracellular juxtamembrane residues.
Purpose of the Study:
- To investigate the biological significance of CSF-1 ectodomain cleavage.
- To compare the biosynthesis and biological activities of uncleavable CSF-1 mutants with wild-type (WT) CSF-1.
Main Methods:
- Generation of uncleavable CSF-1 mutant forms by targeted deletion of extracellular juxtamembrane residues.
- Comparison of cell surface accumulation levels between mutant and WT CSF-1.
- Assessment of biological activities, including CSF-1-dependent cell proliferation and intercellular adhesion.
Main Results:
- Uncleavable CSF-1 forms accumulated at approximately threefold higher levels on the cell surface compared to WT CSF-1.
- Membrane-anchored uncleavable CSF-1 forms were biologically active in promoting proliferation of CSF-1-dependent cells.
- Uncleavable CSF-1 forms exhibited approximately twofold stronger adhesive activity than WT CSF-1.
Conclusions:
- Ectodomain cleavage plays a crucial role in regulating the biological activities of membrane-anchored CSF-1.
- The cleavage process modulates CSF-1's ability to mediate cell proliferation and adhesion.
- Understanding this cleavage mechanism is important for comprehending CSF-1 signaling pathways.