SolubleN-ethylmaleimide-sensitive factor attachment protein receptors required duringTrypanosoma cruziparasitophorous vacuole development

Abstract

Trypanosoma cruzi, the etiologic agent of Chagas disease, is an obligate intracellular parasite that exploits different host vesicular pathways to invade the target cells. Vesicular and target soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are key proteins of the intracellular membrane fusion machinery. During the early times of T. cruzi infection, several vesicles are attracted to the parasite contact sites in the plasma membrane. Fusion of these vesicles promotes the formation of the parasitic vacuole and parasite entry. In this work, we study the requirement and the nature of SNAREs involved in the fusion events that take place during T. cruzi infection. Our results show that inhibition of N-ethylmaleimide-sensitive factor protein, a protein required for SNARE complex disassembly, impairs T. cruzi infection. Both TI-VAMP/VAMP7 and cellubrevin/VAMP3, two v-SNAREs of the endocytic and exocytic pathways, are specifically recruited to the parasitophorous vacuole membrane in a synchronized manner but, although VAMP3 is acquired earlier than VAMP7, impairment of VAMP3 by tetanus neurotoxin fails to reduce T. cruzi infection. In contrast, reduction of VAMP7 activity by expression of VAMP7's longin domain, depletion by small interfering RNA or knockout, significantly decreases T. cruzi infection susceptibility as a result of a minor acquisition of lysosomal components to the parasitic vacuole. In addition, overexpression of the VAMP7 partner Vti1b increases the infection, whereas expression of a KIF5 kinesin mutant reduces VAMP7 recruitment to vacuole and, concomitantly, T. cruzi infection. Altogether, these data support a key role of TI-VAMP/VAMP7 in the fusion events that culminate in the T. cruzi parasitophorous vacuole development.

Autophagy: A necessary process during the Trypanosoma cruzi life-cycle

“…The abolition of this phenomenon, known as reversal infection, by allowing the lysosomal fusion to TcPV, is a key process for the retention of T. cruzi inside the host cell [26]. In agreement with this, further studies from our group demonstrated that the SNARE VAMP7 and its partner Vti1b, both components of the molecular machinery that promotes lysosomal fusion, are required during T. cruzi vacuole development and, as a consequence, for the parasite infection [27]. We have also shown that migration of VAMP7 positive vesicles to the TcPV depends on KIF5, a motor protein belonging to the superfamily of kinesins.…”

Interaction of microbial pathogens with host exocytic pathways

“…Importantly, studies detecting the secretion of lysosomal enzymes or surface localisation of the lysosomal transmembrane protein LAMP1 demonstrated that trypanosomes induce calcium‐mediated exocytosis through fusion of lysosomes with the plasma membrane (Rodriguez, Webster, Ortego, & Andrews, ). Recent data indicate that this fusion event is mediated by VAMP7, a v‐SNARE present on lysosomes (Cueto et al, ). Collectively, these results reveal that T. cruzi mobilises lysosomes, resulting in exocytosis and pathogen uptake into host cells.…”

Molecular interplay of autophagy and endocytosis in human health and diseases

“…After membrane fusion, the soluble NSF protein and soluble αSNAP bind to the inactive cis‐SNARE configuration. NSF then hydrolyses and dissociates the complex so that it is available for the next fusion (Ungar & Hughson, ; Cueto et al , ). STX1, SNAP‐25 and VAMP1 were the first SNAREs to be discovered.…”

“…Vti1p, VAM3p, and VAM7p are vacuolar SNARE proteins, which are the target of the HOPS complex that enhances fusion efficiency in yeast (Wang et al , ). There are nine v‐SNAREs that have been identified in mammals, of which VAMP3 and VAMP7 are involved in endosomal membrane fusion (Cueto et al , ). VAMP3 is present on early and recycling endosomes, and regulates amphisome formation, while VAMP7 is present on late endosomes and post‐Golgi secretory vesicles, and modulates autolysosome formation.…”