Constitutive calcium-independent release of Toxoplasma gondii dense granules occurs through the NSF/SNAP/SNARE/Rab machinery

The signals and the molecular machinery mediating release of dense matrix g ranules from pathogenic protozoan parasites are unknown. We compared the se cretion of the endogenous dense granule marker GRA3 in Toxoplasma gondii wi th the release of a stably transfected foreign reporter, beta-lactamase, th at localizes to parasite dense granules. Both proteins were released consti tutively in a calcium-independent fashion, as shown using both intact and s treptolysin O-permeabilized parasites. N-Ethylmaleimide and recombinant bov ine Rab-guanine dissociation inhibitor inhibited p-lactamase secretion in p ermeabilized parasites, whereas recombinant hamster N-ethylmaleimide-sensit ive fusion protein and bovine alpha-SNAP augmented release. Guanosine 5'-3- O-(thio)triphosphate, but not cAMP, augmented secretion in the presence but not in the absence of ATP. The T. gondii NSF/SNAP/SNARE/Rab machinery part icipates in dense granule release using parasite protein components that ca n interact functionally with their mammalian homologues.