Cytosine Methylation in Genomic DNA and Characterization of DNA Methylases and Demethylases and Their Expression Profiles in Viroid-Infected Hop Plants (Humulus lupulus Var. 'Celeia')

Andrej Sečnik; Nataša Štajner; Sebastjan Radišek; Urban Kunej; Mitja Križman; Jernej Jakše

doi:10.3390/cells11162592

Cytosine Methylation in Genomic DNA and Characterization of DNA Methylases and Demethylases and Their Expression Profiles in Viroid-Infected Hop Plants (Humulus lupulus Var. 'Celeia')

Cells. 2022 Aug 19;11(16):2592. doi: 10.3390/cells11162592.

Authors

Andrej Sečnik¹, Nataša Štajner¹, Sebastjan Radišek², Urban Kunej¹, Mitja Križman³, Jernej Jakše¹

Affiliations

¹ Department of Agronomy, Biotechnical Faculty, University of Ljubljana, 1000 Ljubljana, Slovenia.
² Plant Protection Department, Slovenian Institute of Hop Research and Brewing, 3310 Žalec, Slovenia.
³ Laboratory for Food Chemistry, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana, Slovenia.

Abstract

Abiotic and biotic stresses can lead to changes in host DNA methylation, which in plants is also mediated by an RNA-directed DNA methylation mechanism. Infections with viroids have been shown to affect DNA methylation dynamics in different plant hosts. The aim of our research was to determine the content of 5-methylcytosine (5-mC) in genomic DNA at the whole genome level of hop plants (Humulus lupulus Var. 'Celeia') infected with different viroids and their combinations and to analyse the expression of the selected genes to improve our understanding of DNA methylation dynamics in plant-viroid systems. The adapted HPLC-UV method used proved to be suitable for this purpose, and thus we were able to estimate for the first time that the cytosine methylation level in viroid-free hop plants was 26.7%. Interestingly, the observed 5-mC level was the lowest in hop plants infected simultaneously with CBCVd, HLVd and HSVd (23.7%), whereas the highest level was observed in plants infected with HLVd (31.4%). In addition, we identified three DNA methylases and one DNA demethylase gene in the hop's draft genome. The RT-qPCR revealed upregulation of all newly identified genes in hop plants infected with all three viroids, while no altered expression was observed in any of the other hop plants tested, except for CBCVd-infected hop plants, in which one DNA methylase was also upregulated.

Keywords: 5-mC; DNA demethylase; DNA methylase; DNA methylation; HPLC; RNA-directed DNA methylation; RT-qPCR; hop plant; phylogeny; viroid.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cytosine
DNA
DNA Methylation / genetics
Genomics
Humulus* / genetics
Plant Diseases / genetics
Plants / genetics
Viroids* / genetics

Substances

Cytosine
DNA

Grants and funding

This research was funded by Slovenian Research Agency (SRA-ARRS), grant number P4-0077 Genetics and Modern Technologies of Crops awarded to the research at Chair of Genetics, Biotechnology, Statistics and Plant breeding at the Biotechnical Faculty, University of Ljubljana and young researcher scholarship (200751-53626) awarded to A.S. The APC was funded by grant number P4-0077.