Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins

ABSTRACT Plasmodium parasites must export proteins into their erythrocytic host to survive. Exported proteins must cross the parasite plasma membrane (PPM) and the parasitophorous vacuolar membrane (PVM) encasing the parasite to access the host cell. Crossing the PVM requires protein unfolding and p...

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Autores principales:	Kathryn M. Matthews, Ming Kalanon, Tania F. de Koning-Ward
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	AAA+-ATPase HSP101 PTEX Plasmodium genetic engineering protein trafficking Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/a18b0e57178b4b298b8ddf0b1a9f4091
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spelling	oai:doaj.org-article:a18b0e57178b4b298b8ddf0b1a9f40912021-11-15T15:55:24ZUncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins10.1128/mBio.01106-192150-7511https://doaj.org/article/a18b0e57178b4b298b8ddf0b1a9f40912019-06-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01106-19https://doaj.org/toc/2150-7511ABSTRACT Plasmodium parasites must export proteins into their erythrocytic host to survive. Exported proteins must cross the parasite plasma membrane (PPM) and the parasitophorous vacuolar membrane (PVM) encasing the parasite to access the host cell. Crossing the PVM requires protein unfolding and passage through a translocon, the Plasmodium translocon of exported proteins (PTEX). In this study, we provide the first direct evidence that heat shock protein 101 (HSP101), a core component of PTEX, unfolds proteins for translocation across the PVM by creating transgenic Plasmodium parasites in which the unfoldase and translocation functions of HSP101 have become uncoupled. Strikingly, while these parasites could export native proteins, they were unable to translocate soluble, tightly folded reporter proteins bearing the Plasmodium export element (PEXEL) across the PVM into host erythrocytes under the same conditions. In contrast, an identical PEXEL reporter protein but harboring a transmembrane domain could be exported, suggesting that a prior unfolding step occurs at the PPM. Together, these results demonstrate that the export of parasite proteins is dependent on how these proteins are presented to the secretory pathway before they reach PTEX as well as their folded status. Accordingly, only tightly folded soluble proteins secreted into the vacuolar space and not proteins containing transmembrane domains or the majority of erythrocyte-stage exported proteins have an absolute requirement for the full unfoldase activity of HSP101 to be exported. IMPORTANCE The Plasmodium parasites that cause malaria export hundreds of proteins into their host red blood cell (RBC). These exported proteins drastically alter the structural and functional properties of the RBC and play critical roles in parasite virulence and survival. To access the RBC cytoplasm, parasite proteins must pass through the Plasmodium translocon of exported proteins (PTEX) located at the membrane interfacing the parasite and host cell. Our data provide evidence that HSP101, a component of PTEX, serves to unfold protein cargo requiring translocation. We also reveal that addition of a transmembrane domain to soluble cargo influences its ability to be translocated by parasites in which the HSP101 motor and unfolding activities have become uncoupled. Therefore, we propose that proteins with transmembrane domains use an alternative unfolding pathway prior to PTEX to facilitate export.Kathryn M. MatthewsMing KalanonTania F. de Koning-WardAmerican Society for MicrobiologyarticleAAA+-ATPaseHSP101PTEXPlasmodiumgenetic engineeringprotein traffickingMicrobiologyQR1-502ENmBio, Vol 10, Iss 3 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	AAA+-ATPase HSP101 PTEX Plasmodium genetic engineering protein trafficking Microbiology QR1-502
spellingShingle	AAA+-ATPase HSP101 PTEX Plasmodium genetic engineering protein trafficking Microbiology QR1-502 Kathryn M. Matthews Ming Kalanon Tania F. de Koning-Ward Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins
description	ABSTRACT Plasmodium parasites must export proteins into their erythrocytic host to survive. Exported proteins must cross the parasite plasma membrane (PPM) and the parasitophorous vacuolar membrane (PVM) encasing the parasite to access the host cell. Crossing the PVM requires protein unfolding and passage through a translocon, the Plasmodium translocon of exported proteins (PTEX). In this study, we provide the first direct evidence that heat shock protein 101 (HSP101), a core component of PTEX, unfolds proteins for translocation across the PVM by creating transgenic Plasmodium parasites in which the unfoldase and translocation functions of HSP101 have become uncoupled. Strikingly, while these parasites could export native proteins, they were unable to translocate soluble, tightly folded reporter proteins bearing the Plasmodium export element (PEXEL) across the PVM into host erythrocytes under the same conditions. In contrast, an identical PEXEL reporter protein but harboring a transmembrane domain could be exported, suggesting that a prior unfolding step occurs at the PPM. Together, these results demonstrate that the export of parasite proteins is dependent on how these proteins are presented to the secretory pathway before they reach PTEX as well as their folded status. Accordingly, only tightly folded soluble proteins secreted into the vacuolar space and not proteins containing transmembrane domains or the majority of erythrocyte-stage exported proteins have an absolute requirement for the full unfoldase activity of HSP101 to be exported. IMPORTANCE The Plasmodium parasites that cause malaria export hundreds of proteins into their host red blood cell (RBC). These exported proteins drastically alter the structural and functional properties of the RBC and play critical roles in parasite virulence and survival. To access the RBC cytoplasm, parasite proteins must pass through the Plasmodium translocon of exported proteins (PTEX) located at the membrane interfacing the parasite and host cell. Our data provide evidence that HSP101, a component of PTEX, serves to unfold protein cargo requiring translocation. We also reveal that addition of a transmembrane domain to soluble cargo influences its ability to be translocated by parasites in which the HSP101 motor and unfolding activities have become uncoupled. Therefore, we propose that proteins with transmembrane domains use an alternative unfolding pathway prior to PTEX to facilitate export.
format	article
author	Kathryn M. Matthews Ming Kalanon Tania F. de Koning-Ward
author_facet	Kathryn M. Matthews Ming Kalanon Tania F. de Koning-Ward
author_sort	Kathryn M. Matthews
title	Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins
title_short	Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins
title_full	Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins
title_fullStr	Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins
title_full_unstemmed	Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins
title_sort	uncoupling the threading and unfoldase actions of <italic toggle="yes">plasmodium</italic> hsp101 reveals differences in export between soluble and insoluble proteins
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/a18b0e57178b4b298b8ddf0b1a9f4091
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Uncoupling the Threading and Unfoldase Actions of <italic toggle="yes">Plasmodium</italic> HSP101 Reveals Differences in Export between Soluble and Insoluble Proteins

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