TRAFFIC, MEMBRANE DISTRIBUTION AND SHEDDING OF A VIRULENCE FACTOR OF Trypanosoma cruzi

LANTOS, ANDRES; NIYOGI, SAYANTANEE; ARAOZ, BEATRIZ; CARLEVARO, GIANNINA; BOSSI, MARIANO L.; DOCAMPO, ROBERTO; MUCCI, JUAN; CAMPETELLA, OSCAR

Mar del Plata

Congreso; X Congreso de Protozoología y enfermedades parasitarias (SAP X); 2014

TRAFFIC, MEMBRANE DISTRIBUTION AND SHEDDING OF A VIRULENCE FACTOR OF Trypanosoma cruzi Andrés Lantos(1), Sayantanee Niyogi(2), Beatriz Araoz(3), Giannina Carlevaro(4), Mariano Bossi(5), Roberto Docampo(6), Juan Mucci(7), Oscar Campetella(8) (1)IIB UNSAM. (2)(6)Department of Cellular Biology and Center for Tropical and Emerging Global Diseases and University of Georgia, Athens, Georgia.. (3)(5)INQUIMAE. Departamento de Química Inorgánica, Analítica y Química Física, FCEN-UBA - CONICET. Buenos Aires, Argentina.. (4)(7)(8)Instituto de Investigaciones Biotecnológicas - Universidad Nacional de San Martín - CONICET. Buenos Aires, Argentina.. E-mail: alantos@iibintech.com.ar trans-Sialidase (TS) is a key virulence factor for T. cruzi, the etiological agent of Chagas disease. The enzyme is involved in crucial aspects of the parasite?s life cycle and also generates several abnormalities in the immune system of the infected mammal. TS is a GPI-anchored protein (GPI-AP) located in the plasma membrane of trypomastigotes, which is shed to the milieu. In this work we follow how TS, as well as other GPI-AP like mucins and TSSA are delivered to the plasma membrane via the contractile vacuole complex (CVC) during differentiation of intracellular amastigotes into trypomastigotes. TS is transported from the CVC to the plasma membrane ushered by the small GTPase Rab11. Expression of a dominant negative mutant for Rab11 prevents TS from locating to the plasma membrane and also reduces T. cruzi infection. Infection reduction is rescued by addition of recombinant TS to the culture medium. In the plasma membrane TS appears in discrete domains, which are barely separated one from another when observed under a confocal microscope. To have a better insight of TS distribution we performed super-resolution (GSDIM) imaging of these domains and have determined that TS domains are about 50-nm thick and separated by 100 nm from each other in a zig-zag distribution, validating confocal observations. We evaluated whether TS belongs to detergent resistant domains (DRDs) and found that they do not given that they are susceptible to detergent extraction. However other GPI-AP in trypomastigotes, such as mucins, does belong to DRDs and, correspondingly, does not colocalize with TS in the trypomastigote?s membrane. In epimastigotes, the stage specific mucins (MW 35-50 kDa) also belong to DRD and the traffic to membrane is Rab 11 independent. Lastly, TS is shed to the milieu in exosomes and only appears as a soluble protein after digestion with recombinant PI-PLC. This finding suggests that there is no functional native PI-PLC in T. cruzi trypomastigotes.