Time-gated confocal microscopy reveals accumulation of exocyst subunits at the plant-pathogen interface

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Abstract

Polarized exocytosis is essential for plant development and defence. The exocyst, an octameric protein complex that tethers exocytotic vesicles to the plasma membrane, targets exocytosis. Upon pathogen attack, secreted materials form papillae to halt pathogen penetration. To determine if the exocyst is directly involved in targeting exocytosis to infection sites, information about its localization is instrumental. Here, we investigated exocyst subunit localization in the moss Physcomitrella patens upon pathogen attack and infection by Phytophthora capsici. Time-gated confocal microscopy was used to eliminate autofluorescence of deposited material around infection sites, allowing the visualization of the subcellular localization of exocyst subunits and of v-SNARE Vamp72A1-labelled exocytotic vesicles during infection. This showed that exocyst subunits Sec3a, Sec5b, Sec5d, and Sec6 accumulated at sites of attempted pathogen penetration. Upon pathogen invasion, the exocyst subunits accumulated on the membrane surrounding papilla-like structures and hyphal encasements. Vamp72A1-labelled vesicles were found to localize in the cytoplasm around infection sites. The re-localization of exocyst subunits to infection sites suggests that the exocyst is directly involved in facilitating polarized exocytosis during pathogenesis.
Original languageEnglish
Pages (from-to)837-849
Number of pages13
JournalJournal of Experimental Botany
Volume71
Issue number3
DOIs
Publication statusPublished - 23 Jan 2020

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Confocal Microscopy
plant pathogens
exocytosis
Exocytosis
Infection
pathogens
infection
Bryopsida
material forms
Phytophthora
Bryophyta
SNARE Proteins
Physcomitrella patens
Phytophthora capsici
Plant Development
mosses and liverworts
plant development
confocal microscopy
Cytoplasm
plasma membrane

Cite this

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title = "Time-gated confocal microscopy reveals accumulation of exocyst subunits at the plant-pathogen interface",
abstract = "Polarized exocytosis is essential for plant development and defence. The exocyst, an octameric protein complex that tethers exocytotic vesicles to the plasma membrane, targets exocytosis. Upon pathogen attack, secreted materials form papillae to halt pathogen penetration. To determine if the exocyst is directly involved in targeting exocytosis to infection sites, information about its localization is instrumental. Here, we investigated exocyst subunit localization in the moss Physcomitrella patens upon pathogen attack and infection by Phytophthora capsici. Time-gated confocal microscopy was used to eliminate autofluorescence of deposited material around infection sites, allowing the visualization of the subcellular localization of exocyst subunits and of v-SNARE Vamp72A1-labelled exocytotic vesicles during infection. This showed that exocyst subunits Sec3a, Sec5b, Sec5d, and Sec6 accumulated at sites of attempted pathogen penetration. Upon pathogen invasion, the exocyst subunits accumulated on the membrane surrounding papilla-like structures and hyphal encasements. Vamp72A1-labelled vesicles were found to localize in the cytoplasm around infection sites. The re-localization of exocyst subunits to infection sites suggests that the exocyst is directly involved in facilitating polarized exocytosis during pathogenesis.",
author = "Overdijk, {Elysa J.R.} and Han Tang and Borst, {Jan Willem} and Francine Govers and Tijs Ketelaar",
year = "2020",
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doi = "10.1093/jxb/erz478",
language = "English",
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journal = "Journal of Experimental Botany",
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T1 - Time-gated confocal microscopy reveals accumulation of exocyst subunits at the plant-pathogen interface

AU - Overdijk, Elysa J.R.

AU - Tang, Han

AU - Borst, Jan Willem

AU - Govers, Francine

AU - Ketelaar, Tijs

PY - 2020/1/23

Y1 - 2020/1/23

N2 - Polarized exocytosis is essential for plant development and defence. The exocyst, an octameric protein complex that tethers exocytotic vesicles to the plasma membrane, targets exocytosis. Upon pathogen attack, secreted materials form papillae to halt pathogen penetration. To determine if the exocyst is directly involved in targeting exocytosis to infection sites, information about its localization is instrumental. Here, we investigated exocyst subunit localization in the moss Physcomitrella patens upon pathogen attack and infection by Phytophthora capsici. Time-gated confocal microscopy was used to eliminate autofluorescence of deposited material around infection sites, allowing the visualization of the subcellular localization of exocyst subunits and of v-SNARE Vamp72A1-labelled exocytotic vesicles during infection. This showed that exocyst subunits Sec3a, Sec5b, Sec5d, and Sec6 accumulated at sites of attempted pathogen penetration. Upon pathogen invasion, the exocyst subunits accumulated on the membrane surrounding papilla-like structures and hyphal encasements. Vamp72A1-labelled vesicles were found to localize in the cytoplasm around infection sites. The re-localization of exocyst subunits to infection sites suggests that the exocyst is directly involved in facilitating polarized exocytosis during pathogenesis.

AB - Polarized exocytosis is essential for plant development and defence. The exocyst, an octameric protein complex that tethers exocytotic vesicles to the plasma membrane, targets exocytosis. Upon pathogen attack, secreted materials form papillae to halt pathogen penetration. To determine if the exocyst is directly involved in targeting exocytosis to infection sites, information about its localization is instrumental. Here, we investigated exocyst subunit localization in the moss Physcomitrella patens upon pathogen attack and infection by Phytophthora capsici. Time-gated confocal microscopy was used to eliminate autofluorescence of deposited material around infection sites, allowing the visualization of the subcellular localization of exocyst subunits and of v-SNARE Vamp72A1-labelled exocytotic vesicles during infection. This showed that exocyst subunits Sec3a, Sec5b, Sec5d, and Sec6 accumulated at sites of attempted pathogen penetration. Upon pathogen invasion, the exocyst subunits accumulated on the membrane surrounding papilla-like structures and hyphal encasements. Vamp72A1-labelled vesicles were found to localize in the cytoplasm around infection sites. The re-localization of exocyst subunits to infection sites suggests that the exocyst is directly involved in facilitating polarized exocytosis during pathogenesis.

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DO - 10.1093/jxb/erz478

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