Specific binding of human interferon-gamma to particulates from hemolymph and protocerebrum of tobacco hornworm (Manduca sexta) larvae

Specific binding sites for human interferon-gamma (hIFN-gamma) were found o n particulates prepared from the hemolymph and protocerebrum of fifth-insta r larvae of the tobacco hornworm, Manduca sexta. A portion of these sites c ould be solubilized in an active form by the zwitterionic detergent 3-[(3-c holamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS). A well-defined specific binding was also associated with hemolymph particulate residue af ter solubilization by CHAPS. About one-half of [I-125]hIFN-gamma binding co uld be displaced by heparin. The bound hIFN-gamma could be covalently cross -linked to the binding sites using disuccinylamide suberate, and the molecu lar weight range of these complexes was 200-800 kDa as determined by densit y gradient sedimentation and gel-exclusion chromatography. Only a small fra ction of the hemolymph IFN-gamma binding could be competed by another mamma lian cytokine, rat prolactin (rPRL), while there was no sensitivity to rat growth hormone. The small specific rPRL binding found in Manduca hemolymph showed an affinity similar to the prolactin sites found in the liver of pre gnant rats. The detergent-insoluble Manduca hIFN-gamma binding was bimodal and similar in affinity distribution to the binding found with human platel et membranes (K-diss range 0.1-2 nM). The detergent-solubilized IFN-gamma s ites were homogenous, with a K-diss of about 1.5 nM. The IFN-gamma binding sites in Manduca tissues may therefore include molecular species similar to the known invertebrate cytokine receptors and proteoglycan co-receptors. ( C) 1999 Elsevier Science Inc. All rights reserved.