A novel monomeric adaptor protein in Giardia lamblia with a dual epsin-like function.

FELIZIANI C; ZAMPONI N; MIRAS S; ROPOLO AS; LANFREDI-RANGEL; TOUZ MC

Congreso; Gordon Research Conference. Host-Parasite Interaction; 2012

Endocytosis and lysosomal protein trafficking is essential in pathogenic parasites since it is directly linked to vital parasite-specific processes, e.g. host cell invasion, nutrition, and cell differentiation into resistant stages as in the case of Giardia. The epsin N-terminal homology (ENTH) domain is an evolutionarily conserved protein module found primarily in proteins that participate in clathrin-mediated trafficking. By searching in the GDB, we found the protein GlENTHp (for Giardia lamblia ENTH protein) contains an ENTH domain that shares the 3D structure with the ENTH domain of human epsin, displaying an alpha-helical structure composed of 7 alpha-helices. This domain is present in the epsin or epsin-related (epsinR) form, which are involved in endocytosis and protein trafficking from the Golgi to the lysosomes, respectively. Both epsin and epsinR possess clathrin-binding motifs, but only epsin incorporates an ubiquitin-interaction motif (UIM). Using Giardia?s specific anti-clathrin and anti-ubiquitin antibodies, we determined that GlENTHp interacts with both proteins. Subcellular localization showed this protein mainly in the cytosol and occasionally in the nuclei. Biochemical analysis showed that GlENTHp binds PI(3,4,5)P3 and PI(4)P, phosphoinositides linked to anchoring to the plasma membrane and Golgi, respectively. Moreover, protein-protein interaction experiments showed that GlENTHp associates with the alpha subunit of the adaptor protein-2 (AP2, involved in endocytosis) and the gamma subunit of AP1 (implicated in the Golgi-to-lysosome trafficking). Altogether, these results suggest that GlENTHp participates in the machinery for clathrin-mediated membrane budding acting as a dual epsin-epsinR protein.