Release of the variant surface glycoprotein during differentiation of bloodstream to procyclic forms of Trypanosoma brucei

Investigations on the turnover of the membrane-form variant surface glycoprotein (mfVSG) of Trypanosoma brucei during cultivation in vitro of the monomorphic variant clones MIT at 1.2 and MIT at 1.4 showed that bloodstream forms slowly released the surface coat into the medium (time required to decline to half the initial amount, t50% = 32 +/- 3 h). VSG appeared in the medium in its soluble form (sVSG) which lacked the dimyristoylglycerol membrane anchor as judged by electrophoretic mobility and exposure of the cross-reacting determinant. The total VSG in the culture was very stable with a t50% = 189 +/- 24 h, compared to the other cellular proteins with a t50% approximately 28 h. Coat release during differentiation of bloodstream forms to procyclic cells could be distinguished from this turnover both by its more rapid kinetics (t50% = 13 +/- 1 h) and by the appearance in the medium of a predominant proteolytic fragment in addition to sVSG. Coat release during the transition to procyclic forms was not inhibited by the lysosomotropic agents ammonium chloride or chloroquine, by the proton ionophore monensin, or by the protease inhibitor tosyl-L-lysine chloromethyl ketone. The experiments demonstrate that coat release during differentiation is a specific cellular event distinct from simple turnover. The possibility is discussed that VSG release under both conditions occurs by endocytosis of mfVSG, degradation by a phospholipase C or a protease or both in a non-acidic intracellular compartment and recycling to the surface by exocytosis.