FcgammaRIIA exogenously expressed in HeLa cells activates the mitogen-activated protein kinase cascade by a mechanism dependent on the endogenous expression of the protein tyrosine kinase Syk.
HeLa cells transfected to express the human Fc receptor FcgammaRIIA were stimulated with aggregates of IgG, IgG-ovalbumin equivalence immune complexes and monoclonal antibody reacting with FcgammaRIIA. All of these stimuli activated the cells as judged from the band-shift characteristic of the activation of the p42-MAP/ERK kinase. Since this response is currently associated with the activation of the protein tyrosine kinase Syk, the expression of which is currently thought to be restricted to hemopoietic cells, the results were considered as an indirect evidence of the expression in HeLa cells of either Syk or another protein tyrosine kinase accounting for the same function. Transfection with a dominant negative Syk mutant abrogated the response to FcgammaRIIA cross-linking, whereas overexpression of Syk did not increase the extent of the response. Further evidence of the expression of syk was obtained by the reverse transcription PCR approach and sequencing of the DNA bands. Moreover, immunoprecipitation with anti-Syk antibody of the cell lysates obtained after cross-linking of FcgammaRIIA followed by immunoblotting with anti-phosphotyrosine antibody showed the phosphorylation of a protein band migrating as Syk. These data indicate that expression of FcgammaRIIA on epithelial HeLa cells conveys signals to the p42-MAP/ERK kinase by a mechanism involving the recruitment of Syk. In contrast, cross-linking of this receptor does not yield productive signals coupled to other responses associated to the FcgammaR system such as triggering of the arachidonic acid cascade, activation of the NF-kappaB system and production of chemotactic cytokines.
Renedo
M A
MA
Instituto de Biología y Genética Molecular, Consejo Superior de Investigaciones Científicas, Valladolid, Spain.
Fernández
N
N
Crespo
M S
MS
D000818:Animals; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D016718:Arachidonic Acid / Q000378:metabolism; D018932:Chemokine CCL2 / Q000096:biosynthesis; D003432:Cross-Linking Reagents / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D020672:I-kappa B Proteins; D047908:Intracellular Signaling Peptides and Proteins; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D000072000:NF-KappaB Inhibitor alpha; D010741:Phospholipases A / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011940:Receptor Aggregation; D017452:Receptors, IgG / Q000235:genetics / Q000378:metabolism*; D013552:Swine; D000072377:Syk Kinase; D014162:Transfection
D015703:Antigens, CD; D018932:Chemokine CCL2; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D004792:Enzyme Precursors; C080568:Fc gamma receptor IIA; D020672:I-kappa B Proteins; D047908:Intracellular Signaling Peptides and Proteins; C000606204:NFKBIA protein, human; D017452:Receptors, IgG; D000072000:NF-KappaB Inhibitor alpha; D016718:Arachidonic Acid; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; D019950:Mitogen-Activated Protein Kinase 1; D010741:Phospholipases A