Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections

ABSTRACT Palladin is an important component of motile actin-rich structures and nucleates branched actin filament arrays in vitro. Here we examine the role of palladin during Listeria monocytogenes infections in order to tease out novel functions of palladin. We show that palladin is co-opted by L. ...

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Autores principales:	Aaron S. Dhanda, A. Wayne Vogl, Sharifah E. Albraiki, Carol A. Otey, Moriah R. Beck, Julian A. Guttman
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2018
Materias:	actin nucleation actin polymerization Listeria monocytogenes Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/1e0000c0b2b841b0989e1733dd5add48
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spelling	oai:doaj.org-article:1e0000c0b2b841b0989e1733dd5add482021-11-15T15:53:26ZPalladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections10.1128/mBio.02259-172150-7511https://doaj.org/article/1e0000c0b2b841b0989e1733dd5add482018-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02259-17https://doaj.org/toc/2150-7511ABSTRACT Palladin is an important component of motile actin-rich structures and nucleates branched actin filament arrays in vitro. Here we examine the role of palladin during Listeria monocytogenes infections in order to tease out novel functions of palladin. We show that palladin is co-opted by L. monocytogenes during its cellular entry and intracellular motility. Depletion of palladin resulted in shorter and misshapen comet tails, and when actin- or VASP-binding mutants of palladin were overexpressed in cells, comet tails disintegrated or became thinner. Comet tail thinning resulted in parallel actin bundles within the structures. To determine whether palladin could compensate for the Arp2/3 complex, we overexpressed palladin in cells treated with the Arp2/3 inhibitor CK-666. In treated cells, bacterial motility could be initiated and maintained when levels of palladin were increased. To confirm these findings, we utilized a cell line depleted of multiple Arp2/3 complex subunits. Within these cells, L. monocytogenes failed to generate comet tails. When palladin was overexpressed in this Arp2/3 functionally null cell line, the ability of L. monocytogenes to generate comet tails was restored. Using purified protein components, we demonstrate that L. monocytogenes actin clouds and comet tails can be generated (in a cell-free system) by palladin in the absence of the Arp2/3 complex. Collectively, our results demonstrate that palladin can functionally replace the Arp2/3 complex during bacterial actin-based motility. IMPORTANCE Structures containing branched actin filaments require the Arp2/3 complex. One of the most commonly used systems to study intracellular movement generated by Arp2/3-based actin motility exploits actin-rich comet tails made by Listeria. Using these infections together with live imaging and cell-free protein reconstitution experiments, we show that another protein, palladin, can be used in place of Arp2/3 to form actin-rich structures. Additionally, we show that palladin is needed for the structural integrity of comet tails as its depletion or mutation of critical regions causes dramatic changes to comet tail organization. These findings are the first to identify a protein that can functionally replace the Arp2/3 complex and have implications for all actin-based structures thought to exclusively use that complex.Aaron S. DhandaA. Wayne VoglSharifah E. AlbraikiCarol A. OteyMoriah R. BeckJulian A. GuttmanAmerican Society for Microbiologyarticleactin nucleationactin polymerizationListeria monocytogenesMicrobiologyQR1-502ENmBio, Vol 9, Iss 2 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	actin nucleation actin polymerization Listeria monocytogenes Microbiology QR1-502
spellingShingle	actin nucleation actin polymerization Listeria monocytogenes Microbiology QR1-502 Aaron S. Dhanda A. Wayne Vogl Sharifah E. Albraiki Carol A. Otey Moriah R. Beck Julian A. Guttman Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections
description	ABSTRACT Palladin is an important component of motile actin-rich structures and nucleates branched actin filament arrays in vitro. Here we examine the role of palladin during Listeria monocytogenes infections in order to tease out novel functions of palladin. We show that palladin is co-opted by L. monocytogenes during its cellular entry and intracellular motility. Depletion of palladin resulted in shorter and misshapen comet tails, and when actin- or VASP-binding mutants of palladin were overexpressed in cells, comet tails disintegrated or became thinner. Comet tail thinning resulted in parallel actin bundles within the structures. To determine whether palladin could compensate for the Arp2/3 complex, we overexpressed palladin in cells treated with the Arp2/3 inhibitor CK-666. In treated cells, bacterial motility could be initiated and maintained when levels of palladin were increased. To confirm these findings, we utilized a cell line depleted of multiple Arp2/3 complex subunits. Within these cells, L. monocytogenes failed to generate comet tails. When palladin was overexpressed in this Arp2/3 functionally null cell line, the ability of L. monocytogenes to generate comet tails was restored. Using purified protein components, we demonstrate that L. monocytogenes actin clouds and comet tails can be generated (in a cell-free system) by palladin in the absence of the Arp2/3 complex. Collectively, our results demonstrate that palladin can functionally replace the Arp2/3 complex during bacterial actin-based motility. IMPORTANCE Structures containing branched actin filaments require the Arp2/3 complex. One of the most commonly used systems to study intracellular movement generated by Arp2/3-based actin motility exploits actin-rich comet tails made by Listeria. Using these infections together with live imaging and cell-free protein reconstitution experiments, we show that another protein, palladin, can be used in place of Arp2/3 to form actin-rich structures. Additionally, we show that palladin is needed for the structural integrity of comet tails as its depletion or mutation of critical regions causes dramatic changes to comet tail organization. These findings are the first to identify a protein that can functionally replace the Arp2/3 complex and have implications for all actin-based structures thought to exclusively use that complex.
format	article
author	Aaron S. Dhanda A. Wayne Vogl Sharifah E. Albraiki Carol A. Otey Moriah R. Beck Julian A. Guttman
author_facet	Aaron S. Dhanda A. Wayne Vogl Sharifah E. Albraiki Carol A. Otey Moriah R. Beck Julian A. Guttman
author_sort	Aaron S. Dhanda
title	Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections
title_short	Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections
title_full	Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections
title_fullStr	Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections
title_full_unstemmed	Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections
title_sort	palladin compensates for the arp2/3 complex and supports actin structures during <italic toggle="yes">listeria</italic> infections
publisher	American Society for Microbiology
publishDate	2018
url	https://doaj.org/article/1e0000c0b2b841b0989e1733dd5add48
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Palladin Compensates for the Arp2/3 Complex and Supports Actin Structures during <italic toggle="yes">Listeria</italic> Infections

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