Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion

ABSTRACT Encystation 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,...

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Autores principales:	Jana Krtková, Elizabeth B. Thomas, Germain C. M. Alas, Elisabeth M. Schraner, Habib R. Behjatnia, Adrian B. Hehl, Alexander R. Paredez
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Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/380c2eb7232b48af82d2aad7845cb132
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spelling	oai:doaj.org-article:380c2eb7232b48af82d2aad7845cb1322021-11-15T15:50:19ZRac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion10.1128/mBio.01003-162150-7511https://doaj.org/article/380c2eb7232b48af82d2aad7845cb1322016-09-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01003-16https://doaj.org/toc/2150-7511ABSTRACT Encystation 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-RacCA) revealed fewer but larger ESVs than control cells. Consistent with the phenotype of premature maturation of ESVs in HA-RacCA-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. IMPORTANCE The 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.Jana KrtkováElizabeth B. ThomasGermain C. M. AlasElisabeth M. SchranerHabib R. BehjatniaAdrian B. HehlAlexander R. ParedezAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 7, Iss 4 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Jana Krtková Elizabeth B. Thomas Germain C. M. Alas Elisabeth M. Schraner Habib R. Behjatnia Adrian B. Hehl Alexander R. Paredez Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion
description	ABSTRACT Encystation 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-RacCA) revealed fewer but larger ESVs than control cells. Consistent with the phenotype of premature maturation of ESVs in HA-RacCA-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. IMPORTANCE The 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.
format	article
author	Jana Krtková Elizabeth B. Thomas Germain C. M. Alas Elisabeth M. Schraner Habib R. Behjatnia Adrian B. Hehl Alexander R. Paredez
author_facet	Jana Krtková Elizabeth B. Thomas Germain C. M. Alas Elisabeth M. Schraner Habib R. Behjatnia Adrian B. Hehl Alexander R. Paredez
author_sort	Jana Krtková
title	Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion
title_short	Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion
title_full	Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion
title_fullStr	Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion
title_full_unstemmed	Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion
title_sort	rac regulates <named-content content-type="genus-species">giardia lamblia</named-content> encystation by coordinating cyst wall protein trafficking and secretion
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/380c2eb7232b48af82d2aad7845cb132
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Rac Regulates <named-content content-type="genus-species">Giardia lamblia</named-content> Encystation by Coordinating Cyst Wall Protein Trafficking and Secretion

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