Correlative light-environmental scanning electron microscopy of plasma membrane efflux carriers of plant hormone auxin

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F68081731%3A_____%2F21%3A00546239" target="_blank" >RIV/68081731:_____/21:00546239 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216208:11310/21:10438235

Výsledek na webu

<a href="https://www.mdpi.com/2218-273X/11/10/1407" target="_blank" >https://www.mdpi.com/2218-273X/11/10/1407</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.3390/biom11101407" target="_blank" >10.3390/biom11101407</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Correlative light-environmental scanning electron microscopy of plasma membrane efflux carriers of plant hormone auxin

Popis výsledku v původním jazyce

Fluorescence light microscopy provided convincing evidence for the domain organization of plant plasma membrane (PM) proteins. Both peripheral and integral PM proteins show an inhomo-geneous distribution within the PM. However, the size of PM nanodomains and protein clusters is too small to accurately determine their dimensions and nano-organization using routine confocal fluorescence microscopy and super-resolution methods. To overcome this limitation, we have developed a novel correlative light electron microscopy method (CLEM) using total internal reflection fluorescence microscopy (TIRFM) and advanced environmental scanning electron microscopy (A-ESEM). Using this technique, we determined the number of auxin efflux carriers from the PINFORMED (PIN) family (NtPIN3b-GFP) within PM nanodomains of tobacco cell PM ghosts. Protoplasts were attached to coverslips and immunostained with anti-GFP primary antibody and secondary antibody conjugated to fluorochrome and gold nanoparticles. After imaging the nanodomains within the PM with TIRFM, the samples were imaged with A-ESEM without further processing, and quantification of the average number of molecules within the nanodomain was performed. Without requiring any post-fixation and coating procedures, this method allows to study details of the organization of auxin carriers and other plant PM proteins.

Název v anglickém jazyce

Correlative light-environmental scanning electron microscopy of plasma membrane efflux carriers of plant hormone auxin

Popis výsledku anglicky

Fluorescence light microscopy provided convincing evidence for the domain organization of plant plasma membrane (PM) proteins. Both peripheral and integral PM proteins show an inhomo-geneous distribution within the PM. However, the size of PM nanodomains and protein clusters is too small to accurately determine their dimensions and nano-organization using routine confocal fluorescence microscopy and super-resolution methods. To overcome this limitation, we have developed a novel correlative light electron microscopy method (CLEM) using total internal reflection fluorescence microscopy (TIRFM) and advanced environmental scanning electron microscopy (A-ESEM). Using this technique, we determined the number of auxin efflux carriers from the PINFORMED (PIN) family (NtPIN3b-GFP) within PM nanodomains of tobacco cell PM ghosts. Protoplasts were attached to coverslips and immunostained with anti-GFP primary antibody and secondary antibody conjugated to fluorochrome and gold nanoparticles. After imaging the nanodomains within the PM with TIRFM, the samples were imaged with A-ESEM without further processing, and quantification of the average number of molecules within the nanodomain was performed. Without requiring any post-fixation and coating procedures, this method allows to study details of the organization of auxin carriers and other plant PM proteins.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10611 - Plant sciences, botany

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2021

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Biomolecules

ISSN

2218-273X

e-ISSN

2218-273X

Svazek periodika

11

Číslo periodika v rámci svazku

10

Stát vydavatele periodika

CH - Švýcarská konfederace

Počet stran výsledku

13

Strana od-do

1407

Kód UT WoS článku

000712762200001

EID výsledku v databázi Scopus

2-s2.0-85115616208