Functional analysis of a single chain chimeric alpha/beta-granulocyte-macrophage colony-stimulating factor receptor. Importance of a glutamate residue in the transmembrane region
| Autor: | S, Kafert, S, Luther, I, Böll, K, Wagner, A, Ganser, M, Eder |
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| Rok vydání: | 1999 |
| Předmět: |
Integrins
Base Sequence Recombinant Fusion Proteins Molecular Sequence Data Glutamic Acid Granulocyte-Macrophage Colony-Stimulating Factor Membrane Proteins Protein Serine-Threonine Kinases Flow Cytometry Transfection Antibodies Cell Line Mice Receptors Granulocyte-Macrophage Colony-Stimulating Factor Animals Point Mutation Interleukin-3 Cell Division Cell Size |
| Zdroj: | The Journal of biological chemistry. 274(46) |
| ISSN: | 0021-9258 |
| Popis: | To analyze the function of each subunit of the receptor for granulocyte-macrophage colony-stimulating factor (GM-CSF), GMR, we previously generated a single-chain chimeric receptor by fusion of the extracellular and transmembrane domain from the alpha-subunit (alpha-GMR) to the intracellular part of the beta-subunit (beta-GMR) introducing an additional glutamate residue at the fusion site (alpha/beta-GMR). We demonstrated the capacity of alpha/beta-GMR to bind GM-CSF with low affinity and to induce GM-CSF-dependent activation of tyrosine kinase activity and proliferation in transfected Ba/F3 cells. To further compare the functions of wild type and chimeric receptors, we now report that this alpha/beta-GMR is sufficient to mediate morphological changes, expression of alpha(4)- and beta(1)-integrin receptor subunits, and serine-phosphorylation of Akt kinase. To analyze the function of the glutamate residue at the fusion region of alpha/beta-GMR various point mutants changing this amino acid and its position were expressed in Ba/F3 cells. None of these mutants was capable of supporting GM-CSF-dependent proliferation; however, when beta-GMR was coexpressed, GM-CSF mediated short and long term proliferation. Interestingly, some mutants but not alpha/beta-GMR can induce proliferation in the presence of an anti-alpha-GMR antibody. These data demonstrate the significance of a glutamate residue in the transmembrane region of alpha/beta-GMR for ligand-induced receptor activation. |
| Databáze: | OpenAIRE |
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