Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility

ABSTRACT Invasion of the colon wall by Entamoeba histolytica during amoebic dysentery entails migration of trophozoites through tissue layers that are rich in extracellular matrix. Transcriptional silencing of the E. histolytica surface metalloprotease EhMSP-1 produces hyperadherent less-motile trop...

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Autores principales:	Muhammad M. Hasan, José E. Teixeira, Ying-Wai Lam, Christopher D. Huston
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	ADF/cofilin coactosin Entamoeba histolytica amoebiasis cell motility phosphoproteomics Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/cde7775a75114ada93f1cfd3282f8755
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spelling	oai:doaj.org-article:cde7775a75114ada93f1cfd3282f87552021-11-15T15:56:44ZCoactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility10.1128/mBio.00660-202150-7511https://doaj.org/article/cde7775a75114ada93f1cfd3282f87552020-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00660-20https://doaj.org/toc/2150-7511ABSTRACT Invasion of the colon wall by Entamoeba histolytica during amoebic dysentery entails migration of trophozoites through tissue layers that are rich in extracellular matrix. Transcriptional silencing of the E. histolytica surface metalloprotease EhMSP-1 produces hyperadherent less-motile trophozoites that are deficient in forming invadosomes. Reversible protein phosphorylation is often implicated in regulation of cell motility and invadosome formation. To identify such intermediaries of the EhMSP-1-silenced phenotype, here we compared the phosphoproteomes of EhMSP-1-silenced and vector control trophozoites by using quantitative tandem mass spectrometry-based proteomics. Six proteins were found to be differentially phosphorylated in EhMSP-1-silenced and control cells, including EhCoactosin, a member of the ADF/cofilin family of actin-binding proteins, which was more frequently phosphorylated at serine 147. Regulated overexpression of wild-type, phosphomimetic, and nonphosphorylatable EhCoactosin variants was used to test if phosphorylation functions in control of E. histolytica actin dynamics. Each of the overexpressed proteins colocalized with F-actin during E. histolytica phagocytosis. Nonetheless, trophozoites overexpressing an EhCoactosin phosphomimetic mutant formed more and poorly coordinated cell membrane protrusions compared to those in control or cells expressing a nonphosphorylatable mutant, while trophozoites overexpressing nonphosphorylatable EhCoactosin were significantly more motile within a model of mammalian extracellular matrix. Therefore, although EhCoactosin’s actin-binding ability appeared unaffected by phosphorylation, EhCoactosin phosphorylation helps to regulate amoebic motility. These data help to understand the mechanisms underlying altered adherence and motility in EhMSP-1-silenced trophozoites and lay the groundwork for identifying kinases and phosphatases critical for control of amoebic invasiveness. IMPORTANCE Invasive amoebiasis, caused by the intestinal parasite Entamoeba histolytica, causes life-threatening diarrhea and liver abscesses, but, for unknown reasons, only approximately 10% of E. histolytica infections become symptomatic. A key requirement of invasion is the ability of the parasite to migrate through tissue layers. Here, we systematically looked for differences in protein phosphorylation between control parasites and a previously identified hyperadherent E. histolytica cell line that has reduced motility. We identified EhCoactosin, an actin-binding protein not previously known to be phosphoregulated, as one of the differentially phosphorylated proteins in E. histolytica and demonstrated that EhCoactosin phosphorylation functions in control of cell membrane dynamics and amoebic motility. This and the additional differentially phosphorylated proteins reported lay the groundwork for identifying kinases and phosphatases that regulate tissue invasiveness.Muhammad M. HasanJosé E. TeixeiraYing-Wai LamChristopher D. HustonAmerican Society for MicrobiologyarticleADF/cofilincoactosinEntamoeba histolyticaamoebiasiscell motilityphosphoproteomicsMicrobiologyQR1-502ENmBio, Vol 11, Iss 4 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	ADF/cofilin coactosin Entamoeba histolytica amoebiasis cell motility phosphoproteomics Microbiology QR1-502
spellingShingle	ADF/cofilin coactosin Entamoeba histolytica amoebiasis cell motility phosphoproteomics Microbiology QR1-502 Muhammad M. Hasan José E. Teixeira Ying-Wai Lam Christopher D. Huston Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility
description	ABSTRACT Invasion of the colon wall by Entamoeba histolytica during amoebic dysentery entails migration of trophozoites through tissue layers that are rich in extracellular matrix. Transcriptional silencing of the E. histolytica surface metalloprotease EhMSP-1 produces hyperadherent less-motile trophozoites that are deficient in forming invadosomes. Reversible protein phosphorylation is often implicated in regulation of cell motility and invadosome formation. To identify such intermediaries of the EhMSP-1-silenced phenotype, here we compared the phosphoproteomes of EhMSP-1-silenced and vector control trophozoites by using quantitative tandem mass spectrometry-based proteomics. Six proteins were found to be differentially phosphorylated in EhMSP-1-silenced and control cells, including EhCoactosin, a member of the ADF/cofilin family of actin-binding proteins, which was more frequently phosphorylated at serine 147. Regulated overexpression of wild-type, phosphomimetic, and nonphosphorylatable EhCoactosin variants was used to test if phosphorylation functions in control of E. histolytica actin dynamics. Each of the overexpressed proteins colocalized with F-actin during E. histolytica phagocytosis. Nonetheless, trophozoites overexpressing an EhCoactosin phosphomimetic mutant formed more and poorly coordinated cell membrane protrusions compared to those in control or cells expressing a nonphosphorylatable mutant, while trophozoites overexpressing nonphosphorylatable EhCoactosin were significantly more motile within a model of mammalian extracellular matrix. Therefore, although EhCoactosin’s actin-binding ability appeared unaffected by phosphorylation, EhCoactosin phosphorylation helps to regulate amoebic motility. These data help to understand the mechanisms underlying altered adherence and motility in EhMSP-1-silenced trophozoites and lay the groundwork for identifying kinases and phosphatases critical for control of amoebic invasiveness. IMPORTANCE Invasive amoebiasis, caused by the intestinal parasite Entamoeba histolytica, causes life-threatening diarrhea and liver abscesses, but, for unknown reasons, only approximately 10% of E. histolytica infections become symptomatic. A key requirement of invasion is the ability of the parasite to migrate through tissue layers. Here, we systematically looked for differences in protein phosphorylation between control parasites and a previously identified hyperadherent E. histolytica cell line that has reduced motility. We identified EhCoactosin, an actin-binding protein not previously known to be phosphoregulated, as one of the differentially phosphorylated proteins in E. histolytica and demonstrated that EhCoactosin phosphorylation functions in control of cell membrane dynamics and amoebic motility. This and the additional differentially phosphorylated proteins reported lay the groundwork for identifying kinases and phosphatases that regulate tissue invasiveness.
format	article
author	Muhammad M. Hasan José E. Teixeira Ying-Wai Lam Christopher D. Huston
author_facet	Muhammad M. Hasan José E. Teixeira Ying-Wai Lam Christopher D. Huston
author_sort	Muhammad M. Hasan
title	Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility
title_short	Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility
title_full	Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility
title_fullStr	Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility
title_full_unstemmed	Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility
title_sort	coactosin phosphorylation controls <named-content content-type="genus-species">entamoeba histolytica</named-content> cell membrane protrusions and cell motility
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/cde7775a75114ada93f1cfd3282f8755
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Coactosin Phosphorylation Controls <named-content content-type="genus-species">Entamoeba histolytica</named-content> Cell Membrane Protrusions and Cell Motility

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