Role of clathrin-mediated endocytosis in Giardia lamblia encystation

FELIZIANI C; RIVERO MR; ROPOLO A; TOUZ MC

Congreso; XXXII Reunión Anual de la Sociedad Argentina de Protozoología; 2020

Sociedad Argentina de Protozoología

One way that cells have to exchange material and information with their environment is through cell surface receptors involved in processes that range from nutrient uptake to signaling responses. Consequently, endocytosis constitutes a powerful mechanism to regulate both events. During growth, the protozoa parasite G. lamblia acquires cholesterol from the environment through receptor-mediated endocytosis of LDL lipoprotein and chylomicrons. However, during encystation, there is a decrease in available cholesterol that triggers the differentiation from trophozoites to cysts. In this work, we reveal the involvement of the endocytic machinery in the cell differentiation process of G. lamblia to its resistant form, the cyst. By using wild-type as well as stable negative of knock-down strains for the clathrin adaptors GlAP2 (for G. lamblia Adaptor Protein 2) and GlENTHp (for G. lamblia ENTH protein), immunofluorescence assays, and confocal microscopy, we found that GlAP2 colocalize with ESV (for Encystation Specific vesicles) while the GlENTHp seems not to be colocalized with those vesicles. Moreover, GlAP2 knock-down trophozoites were unable to complete the differentiation process, while GlENTHp knock-down trophozoites accomplished the encystation similarly to wild-type cells. When the number of viable cysts was determined by counting with a haemocytometer, we observed a decrease in cyst production only in GlAP2 knock-down mutants. Also, different expression patterns of GlAP2 and GlENTHp were found during encystation when their mRNAs were analyzed by microarrays. Our results support the hypothesis that it is not the endocytosis mechanism per se that is involved in the generation of cysts but rather the role of GlAP2 in the transport of cyst wall proteins to the surface, for cyst development. However, the restricted localization of GlAP2 may also be important in endocytosis of cyst wall molecules during post-release membrane recycling.