THE CHEMOKINE-C10 - IMMUNOLOGICAL AND FUNCTIONAL-ANALYSIS OF THE SEQUENCE ENCODED BY THE NOVEL 2ND-EXON

The gene for C10, a member of the beta-chemokine family of cytokines, contains a novel second exon encoding a 16 amino acid sequence which i s inserted into the amino-terminal region of the protein, We propose a model of C10 structure in which the sequence encoded by the second ex on is located on the surface of the protein where it interacts with re ceptors and defines the predominant epitope of the C10 protein, To tes t this model we produced C10 protein without the sequence encoded by t he second exon, termed C10(-). Antisera generated against purified C10 recognized C10 protein but reacted only weakly with C10(-) protein, R ecognition of C10 protein by C10 antisera was not blocked by pre-incub ation with a peptide encoding the C10 second exon, indicating that the predominant epitope was not simply the amino acid sequence encoded by the second exon, Antibodies generated against a peptide encoding the C10 second exon sequence recognized C10 on Western blots; this reactio n was competed both by C10 second exon peptide and by C10 protein, Fun ctional studies demonstrated that both C10/FLAG and C10(-)/FLAG elicit chemotaxis of mouse PECs and human PBMCs, but that C10(-)/FLAG appear s to be more potent than C10/FLAG, Thus these data are consistent with our model of C10 in which the sequence encoded by the second exon is located on the surface, participates in defining the predominant epito pe, and plays a key role in recognizing or activating chemokine recept ors. (C) 1996 Academic Press Limited