BFA-sensitive and insensitive exocytic pathways in Entamoeba histolytica trophozoites: their relationship to pathogenesis

Manning‐Cela, Rebeca; Marquez, Claudia; Franco, Elizabeth; Talamás‐Rohana, Patricia; Meza, Isaura

doi:10.1046/j.1462-5822.2003.00332.x

Cited by 36 publications

(32 citation statements)

References 60 publications

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“…Although previous studies had detected antigens using antibodies against GA proteins such as €COP and ARF in vesicles of trophozoites that could be disrupted by Brefeldin A, GA functions per se had not (3,12). We now demonstrate that Eh has GA transporters for UDP-galactose and UDP-glucose.…”

Section: Discussionmentioning

confidence: 58%

“…Cryofixation and cryosubstitution studies showed flattened cisternae in the cytoplasm that resemble GA of higher eukaryotes (11). Antibodies against mammalian GA proteins such as €COP and ARF labeled vesicles of trophozoites that could be disrupted by Brefeldin A and okadaic acid, both known disrupters of GA structures of higher eukaryotes (3,12). Previously, a combination of cytochemical and transmission electron microscopy studies identified Eh vesicles with glucose-6-phosphatase and thiamine pyrophosphatase activities as well as vesicles fluorescently labeled with C6 -12-(N-methyl-N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl))-ceramide.…”

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confidence: 99%

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Golgi and Endoplasmic Reticulum Functions Take Place in Different Subcellular Compartments of Entamoeba histolytica

Bredeston

Caffaro

Samuelson

et al. 2005

Journal of Biological Chemistry

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Entamoeba histolytica is a protozoan parasite that causes dysentery in developing countries of Africa, Asia, and Latin America. The lack of a defined Golgi apparatus in E. histolytica as well as in other protists led to the hypothesis that they had evolved prior to the acquisition of such organelle even though glycoproteins, glycolipids, and antigens have been detected, the latter of which react with antibodies against Golgi apparatus proteins of higher eukaryotes. We here provide direct evidence for Golgi apparatus-like functions in E. histolytica as well as for components of glycoprotein folding quality control. Using a combination of bioinformatic, cell biological, and biochemical approaches we have (a) cloned and expressed the E. histolytica UDP-galactose transporter in Saccharomyces cerevisiae; its K m for UDP-galactose is 2.9 M; (b) characterized vesicles in an extract of the above protist, which transport UDPgalactose into their lumen with a K m of 2.7 M; (c) detected galactosyltransferase activity(ies) in the lumen of the above vesicles with the K m for UDP-galactose, using endogenous acceptors, being 93 M; (d) measured latent apyrase activities in the above vesicles, suggesting they are in the lumen; (e) characterized UDP-glucose transport activities in Golgi apparatus and endoplasmic reticulum-like vesicles with K m s for UDP-glucose of ϳ2-4 M. Although the endoplasmic reticulum-like fraction showed UDP-glucose: glycoprotein glucosyltransferase activity, the Golgi apparatus-like fraction did not. This fraction contained other glucosyltransferases. Together, these studies demonstrate that E. histolytica has different vesicles that play a role in protein glycosylation and folding quality control, analogous to the above organellar functions of higher eukaryotes.

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Section: Discussionmentioning

confidence: 58%

mentioning

confidence: 99%

Golgi and Endoplasmic Reticulum Functions Take Place in Different Subcellular Compartments of Entamoeba histolytica

Bredeston

Caffaro

Samuelson

et al. 2005

Journal of Biological Chemistry

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“…Interestingly, it has been shown that export of newly synthesized proteins to the surface via secretory pathway is not actin-dependent, whereas trafficking in endocytic pathways is actin-dependent in Trypanosoma brucei (32). Although the secretory/endocytic pathway is not well characterized in Entamoeba histolytica, it has been shown that fibronectin receptor and Gal/GalNAc lectin trafficking to plasma membrane is brefeldin-A-dependent, whereas thiolproteinase secretion is brefeldin-A-independent in this parasite (33). Further characterization reveals that overexpression of a GTP-locked mutant of EhRabA alters ER morphology and mislocalizes Gal/GalNAc lectin without affecting the localization of other cell surface proteins, indicating the specific role of EhRabA in early secretory pathway (34).…”

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confidence: 99%

Functional Characterization of Monomeric GTPase Rab1 in the Secretory Pathway of Leishmania

Bahl

Parashar

Malhotra

et al. 2015

Journal of Biological Chemistry

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Leishmania secretes a large number of its effectors to the extracellular milieu. However, regulation of the secretory pathway in Leishmania is not well characterized. Here, we report the cloning, expression, and characterization of the Rab1 homologue from Leishmania. We have found that LdRab1 localizes in Golgi in Leishmania. To understand the role of LdRab1 in the secretory pathway of Leishmania, we have generated transgenic parasites overexpressing GFP-LdRab1:WT, GFP-LdRab1:Q67L (a GTPase-deficient dominant positive mutant of Rab1), and GFP-LdRab1:S22N (a GDP-locked dominant negative mutant of Rab1). Surprisingly, our results have shown that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N does not disrupt the trafficking and localization of hemoglobin receptor in Leishmania. To determine whether the Rab1-dependent secretory pathway is conserved in parasites, we have analyzed the role of LdRab1 in the secretion of secretory acid phosphatase and Ldgp63 in Leishmania. Our results have shown that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N significantly inhibits the secretion of secretory acid phosphatase by Leishmania. We have also found that overexpression of GFP-LdRab1: Q67L or GFP-LdRab1:S22N retains RFP-Ldgp63 in Golgi and blocks the secretion of Ldgp63, whereas the trafficking of RFPLdgp63 in GFP-LdRab1:WT-expressing cells is unaltered in comparison with control cells. Taken together, our results have shown that the Rab1-regulated secretory pathway is well conserved, and hemoglobin receptor trafficking follows an Rab1-independent secretory pathway in Leishmania.Small GTPases of the Rab family are master regulators of intracellular trafficking (1). These proteins are localized on specific compartments and regulate the transport through fusion between two compartments in a nucleotide-dependent process (2, 3). About 70 Rab proteins are identified in mammalian cells. In the endocytic pathway, Rab5 is present in the sorting endosome, whereas Rab4 and Rab11 are localized in recycling endosomes (4, 5). Rab7, Rab9, and Rab24 are associated with the late endosomal compartment. Rab7 mediates transport from the late endosomes to lysosomes, whereas Rab9 regulates the trafficking of lysosomal enzymes from the trans-Golgi network to lysosomes (6 -9). In the secretory pathway, Rab1 regulates the anterograde transport from the endoplasmic reticulum (ER) 3 to the Golgi (10), whereas retrograde transport from Golgi to ER is mediated by Rab2 (11). Rab6 localizes in the Golgi and is involved in intra-Golgi trafficking (12). Moreover, Rab18, Rab30, and Rab43 are found to be important for maintaining Golgi structure (13,14). In addition, transport of cargo from the Golgi to the cell surface is regulated by different Rabs. For example, Rab3 plays a role in release of neurotransmitter, Rab27 regulates the trafficking of lytic granules and melanosomes toward the plasma membrane, and Rab8 is involved in regulating the traffic from the trans-Golgi network to the plasma membrane (15). However, the secretory pathway is n...

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“…For example, in mammalian cells, this reagent can block egress of vesicles from the ER (21) and can induce vesicularization of Golgi apparata (11). In E. histolytica, BFA can alter the morphology of a population of Golgi-specific vesicles and inhibit the trafficking of a subset of secretory proteins (15). Interestingly, in E. histolytica, BFA also induces the formation of tubular structures that colocalize with an ER marker, protein disulfide isomerase (15).…”

Section: Vol 8 2009mentioning

confidence: 99%

“…In E. histolytica, BFA can alter the morphology of a population of Golgi-specific vesicles and inhibit the trafficking of a subset of secretory proteins (15). Interestingly, in E. histolytica, BFA also induces the formation of tubular structures that colocalize with an ER marker, protein disulfide isomerase (15). These structures are similar to those observed in the EhRabAQ84L-expressing cell line; therefore, we explored whether EhRabA might regulate vesicle trafficking pathways that are also sensitive to BFA treatment.…”

Section: Vol 8 2009mentioning

confidence: 99%

Overexpression of a Mutant Form of EhRabA, a Unique Rab GTPase of Entamoeba histolytica , Alters Endoplasmic Reticulum Morphology and Localization of the Gal/GalNAc Adherence Lectin

Welter

Temesvari

2009

Eukaryot Cell

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Entamoeba histolytica is a protozoan parasite that causes amoebic dysentery and liver abscess. Vesicle trafficking events, such as phagocytosis and delivery of plasma membrane proteins, have been implicated in pathogenicity. Rab GTPases are proteins whose primary function is to regulate vesicle trafficking; therefore, understanding the function of Rabs in this organism may provide insight into virulence. E. histolytica possesses a number of unique Rabs that exhibit limited homology to host Rabs. In this study we examined the function of one such Rab, EhRabA, by characterizing a mutant overexpressing a constitutively GTP-bound version of the protein. Overexpression of mutant EhRabA resulted in decreased adhesion to and phagocytosis of human red blood cells and in the appearance of large tubular organelles that could be stained with endoplasmic reticulum (ER)-specific but not Golgi complex-specific antibodies. Consistent with the adhesion defect, two subunits of a cell surface adhesin, the galactose/N-acetylgalactosamine lectin, were mislocalized to the novel organelle. A cysteine protease, EhCP2, was also localized to the ER-like compartment in the mutant; however, the localization of two additional cell surface proteins, Igl and SREHP, remained unchanged in the mutant. The phenotype of the mutant could be recapitulated by treatment with brefeldin A, a cellular toxin that disrupts ER-to-Golgi apparatus vesicle traffic. This suggests that EhRabA influences vesicle trafficking pathways that are also sensitive to brefeldin A. Together, the data indicate that EhRabA directly or indirectly influences the morphology of secretory organelles and regulates trafficking of a subset of secretory proteins in E. histolytica.Entamoeba histolytica, a protozoan parasite, is the causative agent of amoebic dysentery and causes liver abscess. It is prevalent in developing countries that cannot prevent its fecal-oral spread and is responsible for considerable global morbidity and mortality (reviewed in reference 12). Infection is acquired by ingestion of the cyst form of the parasite. Excystation occurs in the small intestine, and released amoeboid trophozoites move to, and colonize, the bowel lumen.

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BFA-sensitive and insensitive exocytic pathways in Entamoeba histolytica trophozoites: their relationship to pathogenesis

Cited by 36 publications

References 60 publications

Golgi and Endoplasmic Reticulum Functions Take Place in Different Subcellular Compartments of Entamoeba histolytica

Golgi and Endoplasmic Reticulum Functions Take Place in Different Subcellular Compartments of Entamoeba histolytica

Functional Characterization of Monomeric GTPase Rab1 in the Secretory Pathway of Leishmania

Overexpression of a Mutant Form of EhRabA, a Unique Rab GTPase of Entamoeba histolytica , Alters Endoplasmic Reticulum Morphology and Localization of the Gal/GalNAc Adherence Lectin

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