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Adaptor protein 2 regulates receptor-mediated endocytosis and cyst formation in Giardia lamblia.

ABSTRACT: The parasite Giardia lamblia possesses PVs (peripheral vacuoles) that function as both endosomes and lysosomes and are implicated in the adaptation, differentiation and survival of the parasite in different environments. The mechanisms by which Giardia traffics essential proteins to these organelles and regulates their secretion have important implications in the control of parasite dissemination. In the present study, we describe the participation of the heterotetrameric clathrin-adaptor protein gAP2 (Giardia adaptor protein 2) complex in lysosomal protein trafficking. A specific monoclonal antibody against the medium subunit (gmu2) of gAP2 showed localization of this complex to the PVs, cytoplasm and plasma membrane in the growing trophozoites. gAP2 also co-localized with clathrin in the PVs, suggesting its involvement in endocytosis. Uptake experiments using standard molecules for the study of endocytosis revealed that gAP2 specifically participated in the endocytosis of LDL (low-density lipoprotein). Targeted down-regulation of the gene encoding gmu2 in growing and encysting trophozoites resulted in a large decrease in the amount of cell growth and cyst wall formation, suggesting a distinct mechanism in which gAP2 is directly involved in both endocytosis and vesicular trafficking.

SUBMITTER: Rivero MR

PROVIDER: S-EPMC2861151 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Adaptor protein 2 regulates receptor-mediated endocytosis and cyst formation in Giardia lamblia.

Rivero Maria R MR Vranych Cecilia V CV Bisbal Mariano M Maletto Belkys A BA Ropolo Andrea S AS Touz Maria C MC

The Biochemical journal 20100428 1

The parasite Giardia lamblia possesses PVs (peripheral vacuoles) that function as both endosomes and lysosomes and are implicated in the adaptation, differentiation and survival of the parasite in different environments. The mechanisms by which Giardia traffics essential proteins to these organelles and regulates their secretion have important implications in the control of parasite dissemination. In the present study, we describe the participation of the heterotetrameric clathrin-adaptor protei ...[more]

PMID: 20199400

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Project description:UnlabelledEncystation of the common intestinal parasite Giardia lamblia involves the production, trafficking, and secretion of cyst wall material (CWM). However, the molecular mechanism responsible for the regulation of these sequential processes remains elusive. Here, we examined the role of GlRac, Giardia's sole Rho family GTPase, in the regulation of endomembrane organization and cyst wall protein (CWP) trafficking. Localization studies indicated that GlRac is associated with the endoplasmic reticulum (ER) and the Golgi apparatus-like encystation-specific vesicles (ESVs). Constitutive GlRac signaling increased levels of the ER marker PDI2, induced ER swelling, reduced overall CWP1 production, and promoted the early maturation of ESVs. Quantitative analysis of cells expressing constitutively active hemagglutinin (HA)-tagged GlRac (HA-Rac(CA)) revealed fewer but larger ESVs than control cells. Consistent with the phenotype of premature maturation of ESVs in HA-Rac(CA)-expressing cells, constitutive GlRac signaling resulted in increased CWP1 secretion and, conversely, morpholino depletion of GlRac blocked CWP1 secretion. Wild-type cells unexpectedly secreted large quantities of CWP1 into the medium, and free CWP1 was used cooperatively during cyst formation. These results, in part, could account for the previously reported observation that G. lamblia encysts more efficiently at high cell densities. These studies of GlRac show that it regulates encystation at several levels, and our findings support its coordinating role as a regulator of CWP trafficking and secretion. The central role of GlRac in regulating membrane trafficking and the cytoskeleton, both of which are essential to Giardia parasitism, further suggests its potential as a novel target for drug development to treat giardiasis.ImportanceThe encystation process is crucial for the transmission of giardiasis and the life cycle of many protists. Encystation for Giardia lamblia involves the assembly of a protective cyst wall via sequential production, trafficking, and secretion of cyst wall material. However, the regulatory pathways that coordinate cargo maturation and secretion remain unknown. Here, we asked whether the signaling activities of G. lamblia's single Rho family GTPase, GlRac, might have a regulatory role in the encystation process. We show that GlRac localizes to endomembranes and its signaling activities regulate the production of cyst wall protein 1 (CWP1), the maturation of encystation-specific vesicles (ESVs), and secretion of CWP1. We also show that secreted CWP1 is available for the development of cysts at the population level, a finding that in part could explain why Giardia encystation proceeds more efficiently at high cell densities.

Dataset Information

Adaptor protein 2 regulates receptor-mediated endocytosis and cyst formation in Giardia lamblia.

Publications

Adaptor protein 2 regulates receptor-mediated endocytosis and cyst formation in Giardia lamblia.

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