An accessory role for the diacylglycerol moiety of variable surface glycoprotein of African trypanosomes in the stimulation of bovine monocytes

The membrane-associated form of the variable surface glycoprotein (mfVSG) f rom African trypanosomes is a potent macrophage activator capable of induci ng production of tumor necrosis factor alpha (TNF alpha) in both bovine and murine models. Using a bovine model, we have re-investigated the hypothesi s that the diacylglycerol moiety of the glycosylphosphatodylinositol (GPI) anchor is involved in macrophage activation and might be the actual parasit e toxin. The anchor of the variable surface glycoprotein (VSG) was labeled with H-3-myristic acid and VSG purified in its,membrane-associated form. Th e dimyristylglycerol moiety of the anchor was released by phospholipase C c leavage. Integrity of the anchor and efficiency of cleavage was verified by autoradiography and methanol:hexane extraction. For analysis of biological function, bovine monocytes were used which had been incubated with bovine interferon gamma (primed) or with culture medium (unprimed). The VSG purifi ed in its membrane-associated form was found to stimulate both primed and u nprimed cells to secrete TNF alpha The same preparation from which the dimy ristylglycerol moiety had been cleaved was no longer able to stimulate unpr imed cells but could still stimulate primed cells. Our data indicate that t he presence of the dimyristylglycerol is not an absolute requirement for in duction of TNF alpha production but can substitute for the interferon gamma priming. Therefore, we favor the hypothesis that stimulation of macrophage s to secrete TNF alpha by the mfVSG is mediated by an as yet unknown trigge r moiety and is facilitated by the dimyristylglycerol anchor. (C) 2001 Else vier Science B.V. All rights reserved.