Double-stranded RNA mediates homology-dependant gene silencing of γ-tubulin in the human parasite Entamoeba histolytica

Vayssié, Laurence; Vargas, Miguel; Weber, Christian; Guillén, Nancy

doi:10.1016/j.molbiopara.2004.07.005

Cited by 72 publications

(66 citation statements)

References 30 publications

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“…Rabring7 is a soluble, ring finger-containing protein and also contributes to microtubule-mediated lysosomal movement (Mizuno et al, 2003). E. histolytica apparently lacks these two effectors, which is consistent with the fact that tubulins and microtubules in E. histolytica are significantly different from other organisms (Roy and Lohia, 2004;Vayssie et al, 2004), and may suggest that vesicular trafficking in E. histolytica depends on microtubule-independent Figure 5. Effects of overexpression of EhRab7A, full-length EhVps26 (V26FL), truncated EhVps26 (V26⌬C), or coexpression of EhRab7A and EhVps26 on the total CP activity.…”

Section: Significance Of the Interaction Between Ehrab7a And The Retrsupporting

confidence: 59%

A Retromerlike Complex Is a Novel Rab7 Effector That Is Involved in the Transport of the Virulence Factor Cysteine Protease in the Enteric Protozoan ParasiteEntamoeba histolytica

Nakada‐Tsukui

Saito‐Nakano

Ali

et al. 2005

MBoC

137

173

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Vesicular trafficking plays an important role in a virulence mechanism of the enteric protozoan parasite Entamoeba histolytica as secreted and lysosomal cysteine protease (CP) contributes to both cytolysis of tissues and degradation of internalized host cells. Despite the primary importance of intracellular sorting in pathogenesis, the molecular mechanism of CP trafficking remains largely unknown. In this report we demonstrate that transport of CP is regulated through a specific interaction of Rab7A small GTPase (EhRab7A) with the retromerlike complex. The amoebic retromerlike complex composed of Vps26, Vps29, and Vps35 was identified as EhRab7A-binding proteins. The amoebic retromerlike complex specifically bound to GTP-EhRab7A, but not GDP-EhRab7A through the direct binding via the carboxy terminus of EhVps26. In erythrophagocytosis the retromerlike complex was recruited to prephagosomal vacuoles, the unique preparatory vacuole of digestive enzymes, and later to phagosomes. This dynamism was indistinguishable from that of EhRab7A, and consistent with the premise that the retromerlike complex is involved in the retrograde transport of putative hydrolase receptor(s) from preparatory vacuoles and phagosomes to the Golgi apparatus. EhRab7A overexpression caused enlargement of lysosomes and decrease of the cellular CP activity. The reduced CP activity was restored by the coexpression of EhVps26, implying that the EhRab7A-mediated transport of CP to phagosomes is regulated by the retromerlike complex. INTRODUCTIONRab GTPases play an essential role to regulate intracellular membrane trafficking. The compartmentalization and functions of Rab proteins are modulated at multiple layers of mechanisms including isoprenylation of the carboxy terminus, nucleotide exchange, and binding of specific effector molecules (Stenmark and Olkkonen, 2001;Takai et al., 2001;Tuvim et al., 2001). Among these modulators, effectors play key roles in the control of 7-90 Rab proteins depending on organisms from fission yeast and amoeba to mammals and plants. A variety of effectors have been identified and shown to interact with specific Rab protein. For instance, regulatory secretions of neurotransmitters from neurons or insulin from pancreatic ␤-cells are regulated by the specific interaction of Rab3 with its effectors (Jahn and Sudhof, 1999). At least four proteins, Rabphilin3, Rim1, Rim2, and Noc2, are known to bind Rab3 and recruit cAMP-GEFII, 14 -3-3, and zyxin, respectively, to regulate cAMP responsiveness, phosphoserine-dependent signaling, and the interaction with cytoskeleton (Kotake et al., 1997;Ozaki et al., 2000;Sun et al., 2003). The specificity of the Rab-effector interaction is attributable to primary sequence motifs in only a few cases including exophilins or Slip/Slac2, Rab3, and Rab27 effectors (Izumi et al., 2003). However, the majority of Rab effectors lack recognizable conserved binding motifs. For instance, Rab5 effectors, rabaptin-5, Rabex-5, EEA1, Rabenosyn-5, hVps34/p150, p110␤/p35␣, rabip4Ј, and rabankyrin-5, which a...

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Section: Significance Of the Interaction Between Ehrab7a And The Retrsupporting

confidence: 59%

A Retromerlike Complex Is a Novel Rab7 Effector That Is Involved in the Transport of the Virulence Factor Cysteine Protease in the Enteric Protozoan ParasiteEntamoeba histolytica

Nakada‐Tsukui

Saito‐Nakano

Ali

et al. 2005

MBoC

137

173

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“…In addition to their association with actin assemblies, mammalian Diaphanous proteins mDia1 and mDia3 associate with spindle microtubular assemblies to regulate spindle positioning and proper chromosome alignment to the kinetochore (26,67). In E. histolytica, microtubular assemblies have been found to be both radial (13,41,57) and bipolar spindles (13). Practically nothing is known about the mode of chromosome alignment or segregation on the radial or bipolar assembly in E. histolytica.…”

Section: Resultsmentioning

confidence: 99%

Entamoeba histolytica Encodes Unique Formins, a Subset of Which Regulates DNA Content and Cell Division

Majumder

Lohia

2008

Infect Immun

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The formin family of proteins mediates dynamic changes in actin assembly in eukaryotes, and therefore it is important to understand the function of these proteins in Entamoeba histolytica, where actin forms the major cytoskeletal network. In this study we have identified the formin homologs encoded in the E. histolytica genome based on sequence analysis. Using multiple tools, we have analyzed the primary sequences of the eight E. histolytica formins and discovered three subsets: (i) E. histolytica formin-1 to -3 (Ehformin-1 to -3), (ii) Ehformin-4, and (iii) Ehformin-5 to -8. Two of these subsets (Ehformin-1 to -3 and Ehformin-4) showed significant sequence differences from their closest homologs, while Ehformin-5 to -8 were unique among all known formins. Since Ehformin-1 to -3 showed important sequence differences from Diaphanous-related formins (DRFs), we have studied the functions of Ehformin-1 and -2 in E. histolytica transformants. Like other DRFs, Ehformin-1 and -2 associated with F-actin in response to serum factors, in pseudopodia, in pinocytic and phagocytic vesicles, and at cell division sites. Ehformin-1 and -2 also localized with the microtubular assembly in the nucleus, indicating their involvement in genome segregation. While increased expression of Ehformin-1 and -2 did not affect phagocytosis or motility, it clearly showed an increase in the number of binucleated cells, the number of nuclei in multinucleated cells, and the average DNA content of each nucleus, suggesting that these proteins regulate both mitosis and cytokinesis in E. histolytica.Polymerization of actin into helical filaments is nucleated by different groups of actin-binding proteins, which in turn are controlled by specific signaling molecules in eukaryotic cells (10, 66). Currently, the three known groups of F-actin nucleating factors are the Arp2/3 complex, formins, and spire (10, 21). Formin homology proteins promote rapid assembly of unbranched actin filaments and local reorganization of higherorder cellular structures to execute specified cytoskeletal functions (reviewed in reference 21). These multidomain proteins have many isoforms that are ubiquitously expressed and perform diverse cellular functions in almost all eukaryotic cells, from vertebrates and plants to unicellular protists (25). The defining feature of formin proteins is the formin homology 2 (FH2) domain, which is required for the nucleation and elongation of nascent actin filaments (24,43,46). The FH1 domain, preceding the FH2 domain, is another key region found in most formins and is made up of tandem repeats of proline and other amino acids (24). Polyproline residues in the FH1 domain interact with profilin to recruit assembly-competent actin monomers in the vicinity of the FH2 domain (50, 52, 61). The FH1 domain also interacts with Src homology 3 (SH3) domainor WW domain-containing proteins (17, 59). Thus, the core FH1-FH2 region nucleates unbranched actin filaments, unlike the Arp2/3 complex, which creates branched filaments (21). Other regulatory domains,...

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“…For E. histolytica, the first evidence that expression of dsRNA can trigger downregulation of a target mRNA was obtained in 2004 (34,65), and since then various methods have been developed to implement the RNAi tool in studies of this parasite (reviewed in reference 77), although a direct linkage between dsRNA effects and the RNAi pathway genes has not been forged as yet. Three AGO and one full-length RdRp homologues have been identified in the E. histolytica genome (Table 1), but BLAST searches for potential Dicer homologues revealed only one candidate with a single RNase III domain (1), whereas most Dicers are endowed with two RNase III domains, which form a pseudodimer giving rise to the enzyme's active site (76).…”

Section: Discovery Of Rna Interference and Its Absence In Certain Parmentioning

confidence: 99%

RNA Interference in Protozoan Parasites: Achievements and Challenges

2011

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Protozoan parasites that profoundly affect mankind represent an exceptionally diverse group of organisms, including Plasmodium, Toxoplasma, Entamoeba, Giardia, trypanosomes, and Leishmania. Despite the overwhelming impact of these parasites, there remain many aspects to be discovered about mechanisms of pathogenesis and how these organisms survive in the host. Combined with the ever-increasing availability of sequenced genomes, RNA interference (RNAi), discovered a mere 13 years ago, has enormously facilitated the analysis of gene function, especially in organisms that are not amenable to classical genetic approaches. Here we review the current status of RNAi in studies of parasitic protozoa, with special emphasis on its use as a postgenomic tool.

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Double-stranded RNA mediates homology-dependant gene silencing of γ-tubulin in the human parasite Entamoeba histolytica

Cited by 72 publications

References 30 publications

A Retromerlike Complex Is a Novel Rab7 Effector That Is Involved in the Transport of the Virulence Factor Cysteine Protease in the Enteric Protozoan ParasiteEntamoeba histolytica

A Retromerlike Complex Is a Novel Rab7 Effector That Is Involved in the Transport of the Virulence Factor Cysteine Protease in the Enteric Protozoan ParasiteEntamoeba histolytica

Entamoeba histolytica Encodes Unique Formins, a Subset of Which Regulates DNA Content and Cell Division

RNA Interference in Protozoan Parasites: Achievements and Challenges

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