In-section Click-iT detection and super-resolution CLEM
analysis of nucleolar ultrastructure and replication in plants

J 2024

In-section Click-iT detection and super-resolution CLEM analysis of nucleolar ultrastructure and replication in plants

FRANEK, Michal, Lenka KOPTASIKOVA, Jiri MIKSATKO, David LIEBL, Eliska MACICKOVA et. al.

Základní údaje

Originální název

In-section Click-iT detection and super-resolution CLEM analysis of nucleolar ultrastructure and replication in plants

Autoři

FRANEK, Michal, Lenka KOPTASIKOVA, Jiri MIKSATKO, David LIEBL, Eliska MACICKOVA, Jakub POSPÍŠIL, Milan EŠNER, Martina DVOŘÁČKOVÁ a Jiří FAJKUS

Vydání

Nature Communications, Berlin, Nature, 2024, 2041-1723

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10400 1.4 Chemical sciences

Stát vydavatele

Německo

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 16.600 v roce 2022

Organizační jednotka

Středoevropský technologický institut

DOI

http://dx.doi.org/10.1038/s41467-024-46324-6

UT WoS

001187872900012

Klíčová slova anglicky

FLUORESCENT PROTEINS; 45S RDNA; S-PHASE; CHEMISTRY; LOCALIZATION; ORGANIZATION; SELECTIVITY; FASCIATA1; CELLS; LIGHT

Štítky

CF CELLIM

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 18. 9. 2024 08:58, Mgr. Eva Dubská

Anotace

V originále

Correlative light and electron microscopy (CLEM) is an important tool for the localisation of target molecule(s) and their spatial correlation with the ultrastructural map of subcellular features at the nanometre scale. Adoption of these advanced imaging methods has been limited in plant biology, due to challenges with plant tissue permeability, fluorescence labelling efficiency, indexing of features of interest throughout the complex 3D volume and their re-localization on micrographs of ultrathin cross-sections. Here, we demonstrate an imaging approach based on tissue processing and embedding into methacrylate resin followed by imaging of sections by both, single-molecule localization microscopy and transmission electron microscopy using consecutive CLEM and same-section CLEM correlative workflow. Importantly, we demonstrate that the use of a particular type of embedding resin is not only compatible with single-molecule localization microscopy but shows improvements in the fluorophore blinking behavior relative to the whole-mount approaches. Here, we use a commercially available Click-iT ethynyl-deoxyuridine cell proliferation kit to visualize the DNA replication sites of wild-type Arabidopsis thaliana seedlings, as well as fasciata1 and nucleolin1 plants and apply our in-section CLEM imaging workflow for the analysis of S-phase progression and nucleolar organization in mutant plants with aberrant nucleolar phenotypes.

Návaznosti

EF16_013/0001775, projekt VaV

Název: Modernizace a podpora výzkumných aktivit národní infrastruktury pro biologické a medicínské zobrazování Czech-BioImaging

EF18_046/0016045, projekt VaV

Název: Modernizace národní infrastruktury pro biologické a medicínské zobrazování Czech-BioImaging

GA23-06643S, projekt VaV

Název: Dispozice, výskyt a zpracování DNA lézí v genomu rostlin Arabidopsis thaliana defektních pro reparaci DNA a sestavování nukleozomů

Investor: Grantová agentura ČR, Dispozice, výskyt a zpracování DNA lézí v genomu rostlin Arabidopsis thaliana defektních pro reparaci DNA a sestavování nukleozomů

GX20-01331X, projekt VaV

Název: Biogeneze a evoluce telomerázy u rostlin

Investor: Grantová agentura ČR, Biogenesis and evolution of telomerase in plants

LM2023050, projekt VaV

Název: Národní infrastruktura pro biologické a medicínské zobrazování

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, Czech BioImaging: Národní výzkumná infrastruktura pro biologické a medicínské zobrazování

Citovat

FRANEK, Michal, Lenka KOPTASIKOVA, Jiri MIKSATKO, David LIEBL, Eliska MACICKOVA, Jakub POSPÍŠIL, Milan EŠNER, Martina DVOŘÁČKOVÁ a Jiří FAJKUS. In-section Click-iT detection and super-resolution CLEM analysis of nucleolar ultrastructure and replication in plants. Nature Communications. Berlin: Nature, 2024, roč. 15, č. 1, s. 1-13. ISSN 2041-1723. Dostupné z: https://dx.doi.org/10.1038/s41467-024-46324-6.

@article{2433818,
   author = {Franek, Michal and Koptasikova, Lenka and Miksatko, Jiri and Liebl, David and Macickova, Eliska and Pospíšil, Jakub and Ešner, Milan and Dvořáčková, Martina and Fajkus, Jiří},
   article_location = {Berlin},
   article_number = {1},
   doi = {http://dx.doi.org/10.1038/s41467-024-46324-6},
   keywords = {FLUORESCENT PROTEINS; 45S RDNA; S-PHASE; CHEMISTRY; LOCALIZATION; ORGANIZATION; SELECTIVITY; FASCIATA1; CELLS; LIGHT},
   language = {eng},
   issn = {2041-1723},
   journal = {Nature Communications},
   title = {In-section Click-iT detection and super-resolution CLEM analysis of nucleolar ultrastructure and replication in plants},
   url = {https://www.nature.com/articles/s41467-024-46324-6},
   volume = {15},
   year = {2024}
}

TY  - JOUR
ID  - 2433818
AU  - Franek, Michal - Koptasikova, Lenka - Miksatko, Jiri - Liebl, David - Macickova, Eliska - Pospíšil, Jakub - Ešner, Milan - Dvořáčková, Martina - Fajkus, Jiří
PY  - 2024
TI  - In-section Click-iT detection and super-resolution CLEM analysis of nucleolar ultrastructure and replication in plants
JF  - Nature Communications
VL  - 15
IS  - 1
SP  - 1-13
EP  - 1-13
PB  - Nature
SN  - 20411723
KW  - FLUORESCENT PROTEINS
KW  - 45S RDNA
KW  - S-PHASE
KW  - CHEMISTRY
KW  - LOCALIZATION
KW  - ORGANIZATION
KW  - SELECTIVITY
KW  - FASCIATA1
KW  - CELLS
KW  - LIGHT
UR  - https://www.nature.com/articles/s41467-024-46324-6
N2  - Correlative light and electron microscopy (CLEM) is an important tool for the localisation of target molecule(s) and their spatial correlation with the ultrastructural map of subcellular features at the nanometre scale. Adoption of these advanced imaging methods has been limited in plant biology, due to challenges with plant tissue permeability, fluorescence labelling efficiency, indexing of features of interest throughout the complex 3D volume and their re-localization on micrographs of ultrathin cross-sections. Here, we demonstrate an imaging approach based on tissue processing and embedding into methacrylate resin followed by imaging of sections by both, single-molecule localization microscopy and transmission electron microscopy using consecutive CLEM and same-section CLEM correlative workflow. Importantly, we demonstrate that the use of a particular type of embedding resin is not only compatible with single-molecule localization microscopy but shows improvements in the fluorophore blinking behavior relative to the whole-mount approaches. Here, we use a commercially available Click-iT ethynyl-deoxyuridine cell proliferation kit to visualize the DNA replication sites of wild-type Arabidopsis thaliana seedlings, as well as fasciata1 and nucleolin1 plants and apply our in-section CLEM imaging workflow for the analysis of S-phase progression and nucleolar organization in mutant plants with aberrant nucleolar phenotypes.
ER  -

FRANEK, Michal, Lenka KOPTASIKOVA, Jiri MIKSATKO, David LIEBL, Eliska MACICKOVA, Jakub POSPÍŠIL, Milan EŠNER, Martina DVOŘÁČKOVÁ a Jiří FAJKUS. In-section Click-iT detection and super-resolution CLEM analysis of nucleolar ultrastructure and replication in plants. \textit{Nature Communications}. Berlin: Nature, 2024, roč.~15, č.~1, s.~1-13. ISSN~2041-1723. Dostupné z: https://dx.doi.org/10.1038/s41467-024-46324-6.

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