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SUMMARY Medicago truncatulais a model legume for fundamental research on legume biology and symbiotic nitrogen fixation.Tnt1, a retrotransposon from tobacco, was used to generate insertion mutants inM. truncatulaR108. Approximately 21 000 insertion lines have been generated and publicly available.Tnt1retro‐transposition event occurs during somatic embryogenesis (SE), a pivotal process that triggers massive methylation changes. We studied the SE ofM. truncatulaR108 using leaf explants and explored the dynamic shifts in the methylation landscape from leaf explants to callus formation and finally embryogenesis. Higher cytosine methylation in all three contexts of CG, CHG, and CHH patterns was observed during SE compared to the controls. Higher methylation patterns were observed in assumed promoter regions (~2‐kb upstream regions of transcription start site) of the genes, while lowest was recorded in the untranslated regions. Differentially methylated promoter region analysis showed a higher CHH methylation in embryogenesis tissue samples when compared to CG and CHG methylation. Strong correlation (89.71%) was identified between the differentially methylated regions (DMRs) and the site ofTnt1insertions inM. truncatulaR108 and stronger hypermethylation of genes correlated with higher number ofTnt1insertions in all contexts of CG, CHG, and CHH methylation. Gene ontology enrichment and KEGG pathway enrichment analysis identified genes and pathways enriched in the signal peptide processing, ATP hydrolysis, RNA polymerase activity, transport, secondary metabolites, and nitrogen metabolism pathways. Combined gene expression analysis and methylation profiling showed an inverse relationship between methylation in the DMRs (regions spanning genes) and the expression of genes. Our results show that a dynamic shift in methylation happens during the SE process in the context of CG, CHH and CHG methylation, and theTnt1retrotransposition correlates with the hyperactive methylation regions.
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Genome‐wide loss of CHH methylation with limited transcriptome changes in Setaria viridis DOMAINS REARRANGED METHYLTRANSFERASE ( DRM ) mutants
SUMMARY The DOMAINS REARRANGED METHYLTRANSFERASEs (DRMs) are crucial for RNA‐directed DNA methylation (RdDM) in plant species.Setaria viridisis a model monocot species with a relatively compact genome that has limited transposable element (TE) content. CRISPR‐based genome editing approaches were used to create loss‐of‐function alleles for the two putative functional DRM genes inS. viridisto probe the role of RdDM. Double mutant (drm1ab)plants exhibit some morphological abnormalities but are fully viable. Whole‐genome methylation profiling provided evidence for the widespread loss of methylation in CHH sequence contexts, particularly in regions with high CHH methylation in wild‐type plants. Evidence was also found for the locus‐specific loss of CG and CHG methylation, even in some regions that lack CHH methylation. Transcriptome profiling identified genes with altered expression in thedrm1abmutants. However, the majority of genes with high levels of CHH methylation directly surrounding the transcription start site or in nearby promoter regions in wild‐type plants do not have altered expression in thedrm1abmutant, even when this methylation is lost, suggesting limited regulation of gene expression by RdDM. Detailed analysis of the expression of TEs identified several transposons that are transcriptionally activated indrm1abmutants. These transposons are likely to require active RdDM for the maintenance of transcriptional repression.
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- PAR ID:
- 10445834
- Publisher / Repository:
- Wiley-Blackwell
- Date Published:
- Journal Name:
- The Plant Journal
- Volume:
- 111
- Issue:
- 1
- ISSN:
- 0960-7412
- Page Range / eLocation ID:
- p. 103-116
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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