EhRabB mobilises the EhCPADH complex through the actin cytoskeleton during phagocytosis of
            <i>Entamoeba histolytica</i>

Javier-Reyna, Rosario; Montaño, Sarita; Garcı́a-Rivera, Guillermina; Rodríguez, Mario A.; González‐Robles, Arturo; Orozco, Esther

doi:10.1111/cmi.13071

Cited by 13 publications

(19 citation statements)

References 67 publications

(99 reference statements)

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“…It also appeared around ingested erythrocytes and surrounding big pockets that could be the pre-phagosomes formed to receive the red blood cells from other endosomes ( Figure 7 A). In some images, EhADH did not show up in the same region of the phagocytic channel that EhSUMO did, although laser sections revealed its presence in other places of this structure, as published before [ 38 ]. Large bags in the cytoplasm, that could correspond to putative pre-phagosome structures, were also enlightened by the α-EhADH antibodies, inside and surrounding them, and the label was also detected around the ingested erythrocytes ( Figure 7 A).…”

Section: Resultssupporting

confidence: 71%

Protein Sumoylation Is Crucial for Phagocytosis in Entamoeba histolytica Trophozoites

Díaz-Hernández

Javier-Reyna

Sotto-Ortega

et al. 2021

IJMS

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Posttranslational modifications provide Entamoeba histolytica proteins the timing and signaling to intervene during different processes, such as phagocytosis. However, SUMOylation has not been studied in E. histolytica yet. Here, we characterized the E. histolytica SUMO gene, its product (EhSUMO), and the relevance of SUMOylation in phagocytosis. Our results indicated that EhSUMO has an extended N-terminus that differentiates SUMO from ubiquitin. It also presents the GG residues at the C-terminus and the ΨKXE/D binding motif, both involved in target protein contact. Additionally, the E. histolytica genome possesses the enzymes belonging to the SUMOylation-deSUMOylation machinery. Confocal microscopy assays disclosed a remarkable EhSUMO membrane activity with convoluted and changing structures in trophozoites during erythrophagocytosis. SUMOylated proteins appeared in pseudopodia, phagocytic channels, and around the adhered and ingested erythrocytes. Docking analysis predicted interaction of EhSUMO with EhADH (an ALIX family protein), and immunoprecipitation and immunofluorescence assays revealed that the association increased during phagocytosis; whereas the EhVps32 (a protein of the ESCRT-III complex)-EhSUMO interaction appeared stronger since basal conditions. In EhSUMO knocked-down trophozoites, the bizarre membranous structures disappeared, and EhSUMO interaction with EhADH and EhVps32 diminished. Our results evidenced the presence of a SUMO gene in E. histolytica and the SUMOylation relevance during phagocytosis. This is supported by bioinformatics screening of many other proteins of E. histolytica involved in phagocytosis, which present putative SUMOylation sites and the ΨKXE/D binding motif.

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Section: Resultssupporting

confidence: 71%

Protein Sumoylation Is Crucial for Phagocytosis in Entamoeba histolytica Trophozoites

Díaz-Hernández

Javier-Reyna

Sotto-Ortega

et al. 2021

IJMS

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“…7A). In some images, EhADH did not show up in the same region of the phagocytic channel that EhSUMO did, although laser sections revealed its presence in other places of this structure, as published before (38). Large bags in the cytoplasm, that could correspond to a putative pre-phagosome structures, were also enlightened by the α-EhADH antibodies, inside and surrounding them, and the label was also detected around the ingested erythrocytes (Fig.…”

Section: Resultssupporting

confidence: 77%

Protein SUMOylation is crucial for phagocytosis inEntamoeba histolyticatrophozoites

Díaz-Hernández

Javier-Reyna

Ortega-Soto

et al. 2021

Preprint

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During phagocytosis, a key event in the virulence of the protozoan Entamoeba histolytica , several molecules in concert contact the target, generate pseudopodia, and internalize and digest the ingested prey. Posttranslational modifications provide proteins the timing and signaling to intervene in these processes. SUMOylation is a posttranslational modification that in several systems grants a fine tuning for protein functions, protein interactions and cellular location, but it has not been studied in E. histolytica. In this paper, we characterized the E. histolytica SUMO gene and its product (EhSUMO) and elucidated the EhSUMO 3D-structure. Furthermore, here we studied the relevance of SUMOylation in phagocytosis, particularly in its association with EhADH (an ALIX family protein) and EhVps32 (a protein of the ESCRT-III complex), both involved in phagocytosis. Our results indicated that EhSUMO has an extended N-terminus that differentiates other SUMO from ubiquitin. It also presents the GG residues at the C-terminus and the ΨKXE/D binding motif, both involved in target protein contact. Additionally, E. histolytica genome possesses the enzymes belonging to the SUMOylation-deSUMOylation machineries. Confocal microscopy assays, using a-EhSUMO antibodies disclosed a remarkable membrane activity with convoluted and changing structures in trophozoites during erythrophagocytosis. SUMOylated proteins appeared in pseudopodia, phagocytic channels, and around the adhered and ingested erythrocytes. Docking analysis predicted interaction of EhSUMO with EhADH, and immunoprecipitation and immunofluorescence assays revealed that the EhADH-EhSUMO association increased during phagocytosis, whereas the EhVps32-EhSUMO interaction appeared stronger since basal conditions. In EhSUMO knocked down trophozoites, the bizarre membranous structures disappeared, and EhSUMO interaction with EhADH and EhVps32 diminished. Our results evidenced the presence of a SUMO gene in E. histolytica and the SUMOylation relevance during phagocytosis. Author’s Abstract Phagocytosis is one of the main functions that Entamoeba histolytica trophozoites carry out during the invasion to the host. Many proteins are involved in this fascinating event, in which the plasmatic membrane undergoes to multiple and speedy changes. Posttraductional modifications activate proteins in the precise time that they must get involved. SUMOylation, that consists in the non-covalent binding of SUMO protein with target molecules, is one of the main changes suffered by proteins in order to enable them to participate in cellular functions. SUMOylation had not been studied in E. histolytica nor in phagocytosis, and our working hypothesis is that this event is deeply engaged in the ingestion of target molecules and cells. The results of this paper prove the presence of an intronless bona fide EhSUMO gene encoding for a predicted 12.6 kDa protein that is actively involved in phagocytosis. Silencing of the EhSUMO gene affected the rate of phagocytosis and interfered with the EhADH and EhVps32 function, two proteins involved in phagocytosis, strongly supporting the importance of SUMOylation in this event.

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“…To investigate the functional relevance of EhGATA in E. histolytica , we first, attempted to silence the Ehgata gene expression using the Ehgata ‐dsRNA strategy (Solis et al ., 2009). We PCR‐amplified a 400 bp fragment of the Ehgata gene and cloned it into the pL4440 plasmid to produce dsRNAs, but no Ehgata‐ silenced trophozoites were obtained in our hands, even that using this technique we have successfully silenced other genes (López‐Rosas et al ., 2012; Javier‐Reyna et al ., 2019). In many organisms, GATA proteins exist as families and regulate the expression of genes involved in vital functions.…”

Section: Resultsmentioning

confidence: 99%

“…At 5 min of RBC engulfment, EhADH levels diminished in NeoGATA trophozoites, in comparison to basal condition (Figure 11A,B). In contrast, in Neo trophozoites, the amount of EhADH increases through the phagocytosis kinetics, consistently with earlier findings (Javier‐Reyna et al ., 2019). Meanwhile, EhVps32 augmented in NeoGATA trophozoites after 5 min incubation with RBC, and later it recovered its level as in the basal condition, although it was always in a lower amount than in Neo trophozoites (Figure 11C,D).…”

Section: Resultsmentioning

confidence: 99%

A noncanonical GATA transcription factor of Entamoeba histolytica modulates genes involved in phagocytosis

Huerta¹,

Reyes²,

Garcı́a-Rivera³

et al. 2020

Molecular Microbiology

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EhRabB mobilises the EhCPADH complex through the actin cytoskeleton during phagocytosis of Entamoeba histolytica

Cited by 13 publications

References 67 publications

Protein Sumoylation Is Crucial for Phagocytosis in Entamoeba histolytica Trophozoites

Protein Sumoylation Is Crucial for Phagocytosis in Entamoeba histolytica Trophozoites

Protein SUMOylation is crucial for phagocytosis inEntamoeba histolyticatrophozoites

A noncanonical GATA transcription factor of Entamoeba histolytica modulates genes involved in phagocytosis

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