LUJAN Hugo Daniel
congresos y reuniones científicas
Molecular Mechanims of intracellular protein trafficking during cyst wall formation in Giardia
VANINA ELIANA, ELIAS; RODRIGO QUIROGA; HUGO DANIEL LUJÁN
Congreso; Ibero American Congress on Cell Biology; 2007
Sociedad Iberoamericana de Biologia celular y Universidad Autonoma de Mexico
Giardia trophozoites undergo fundamental changes to survive outside the intestine of their hosts by differentiating into infective cysts. Encystation entails the synthesis, processing, transport, secretion, and assembly of cyst wall materials into a protective cyst wall. We are interested in deciphering the molecular events leading to the formation of the cyst wall, with emphasis on the vesicular transport of cyst wall components. In Giardia there are three closely related proteins that localize within encystation-specific secretory transport vesicles (ESVs) in encysting trophozoites and in the cyst wall of mature cysts. The cyst wall protein genes predict proteins of 26 (CWP1), 39 (CWP2), and 27 (CWP3) kDa, having ~60% identity in a 26 kDa overlapping region. CWP2 differs from CWP1 and CWP3 by a 121-residue carboxy-terminal extension rich in basic amino acids. In this work we (a) determined the role of the alkaline extension of CWP2 in ESV formation, and (b) identified and characterized SNARE proteins involved in Giardia encystation. First, CWP constructs with or without the CWP2 basic tail were transfected into trophozoites and two types mammalian cells, those with and without endogenous secretory granules. Our results demonstrated that the CWP2 basic tail is necessary but not sufficient to induce the biogenesis of ESVs in non-encysting trophozoites and that there must exist a receptor for this tail in Giardia since no granules were generated by expression of these proteins in mammalian cells. After their formation, little is known about the molecular mechanisms involved in vesicular docking and fusion of ESVs in Giardia. In higher eukaryotes, soluble N-ethylmaleimide-sensitive factor-attachment protein receptor (SNARE) of the vesicle-associated membrane proteins (VAMP) and Syntaxin proteins play essential roles in these processes through the formation of complexes between proteins present on donor and target membranes. Here, we also present results from a complete search of the Giardia genome for SNARE encoding genes, the analysis of the structures and subcellular localization of their products, and the functionality of the SNAREs identified in this parasite. Besides, we demonstrate that either the expression or the localization of some of these SNAREs is modified during trophozoite differentiation into cysts. We also found that Giardia SNAREs distinctively localize to different subcellular organelles and that the specificity and degree of interactions between Q-SNAREs and R-SNAREs indicate that Giardia requires a small number of SNAREs to accomplish all the vesicle docking and fusion events that are critical for growth, adaptation, and differentiation.