A role of age-dependent DNA methylation reprogramming in regulating the regeneration capacity of Boea hygrometrica leaves
Plants can regenerate new individuals under appropriate culture conditions. Although the molecular basis of shoot regeneration has steadily been unraveled, the role of age-dependent DNA methylation status in the regulation of explant regeneration remains practically unknown. Here, we established an effective auxin/cytokinin-induced shoot regeneration system for the resurrection plant Boea hygrometrica via direct organogenesis and observed that regeneration was postponed with increasing age of donor plants. Global transcriptome analysis revealed significant upregulation of genes required for hormone signaling and phenylpropanoid biosynthesis and downregulation of photosynthetic genes during regeneration. Transcriptional changes in the positive/negative regulators and cell wall-related proteins involved in plant regeneration, such as ELONGATED HYPOCOTYL5 (HY5), LATERAL ORGAN BOUNDARIES DOMAIN, SHOOT-MERISTEMLESS, and WUSCHEL, were associated with the regeneration process. Comparison of DNA methylation profiling between leaves from young seedlings (YL) and mature plants (ML) revealed increased asymmetrical methylation in ML, which was predominantly distributed in promoter regions of genes, such as HY5 and a member of ABA-responsive element (ABRE) binding protein/ABRE binding factor, as well as genes encoding glycine-rich cell wall structural protein, CENTRORADIALIS-like protein, and beta-glucosidase 40-like essential for shoot meristem and cell wall architecture. Their opposite transcription response in ML explants during regeneration compared with those from YL demonstrated the putative involvement of DNA methylation in regeneration. Moreover, a significant lower expression of DNA glycosylase-lyase required for DNA demethylation in ML was coincident with its postponed regeneration compared with those in YL. Taken together, our results suggest a role of promoter demethylation in B. hygrometrica regeneration.
KeywordsShoot regeneration Donor plant age DNA methylome Transcriptome Resurrection plant Boea hygrometrica
bisulfite sequencing PCR
differentially expressed gene
differentially methylated gene
differentially methylated region
Kyoto Encyclopedia of Genes and Genomes
leaves from mature plants
quantitative real-time PCR
high-throughput mRNA sequencing
whole-genome bisulfite sequencing
leaves from young seedlings
XD conceived the experiments. EZ, JQ, and YL prepared the plant materials. RS analyzed the data and wrote the manuscript. XD and CL revised the manuscript. All authors have read and approved the manuscript.
This work was supported by the National Natural Science Foundation of China (grant numbers 31770293 and 31470361).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
- Berckmans B, Vassileva V, Schmid SPC, Maes S, Parizot B, Naramoto S, Magyar Z, Kamei CLA, Koncz C, Bögre L, Persiau G, de Jaeger G, Friml J, Simon R, Beeckman T, de Veylder L (2011) Auxin-dependent cell cycle reactivation through transcriptional regulation of Arabidopsis E2Fa by lateral organ boundary proteins. Plant Cell 23:3671–3683CrossRefGoogle Scholar
- Iwase A, Harashima H, Ikeuchi M, Rymen B, Ohnuma M, Komaki S, Morohashi K, Kurata T, Nakata M, Ohme-Takagi M, Grotewold E, Sugimoto K (2017) WIND1 promotes shoot regeneration through transcriptional activation of ENHANCER OF SHOOT REGENERATION1 in Arabidopsis. Plant Cell 29:54–69CrossRefGoogle Scholar
- Lister R, Pelizzola M, Dowen RH, Hawkins RD, Hon G, Tonti-Filippini J, Nery JR, Lee L, Ye Z, Ngo QM, Edsall L, Antosiewicz-Bourget J, Stewart R, Ruotti V, Millar AH, Thomson JA, Ren B, Ecker JR (2009) Human DNA methylomes at base resolution show widespread epigenomic differences. Nature 462:315–322CrossRefGoogle Scholar
- Mohebodini M, Mokhtar JJ, Mahboudi F, Alizadeh H (2011) Effects of genotype, explant age and growth regulators on callus induction and direct shoot regeneration of lettuce (Lactuca sativa L.). Aust J Crop Sci 5:92–95Google Scholar
- Nameth B, Dinka SJ, Chatfield SP, Morris A, English J, Lewis D, Oro R, Raizada MN (2013) The shoot regeneration capacity of excised Arabidopsis cotyledons is established during the initial hours after injury and is modulated by a complex genetic network of light signalling. Plant Cell Environ 36:68–86CrossRefGoogle Scholar
- Skoog F, Miller CO (1957) Chemical regulation of growth and organ formation in plant tissues cultured in vitro. Symp Soc Exp Biol 11:118–130Google Scholar
- Sun R-Z, Cheng G, Li Q, He Y-N, Wang Y, Lan Y-B, Li S-Y, Zhu Y-R, Song W-F, Zhang X, Cui X-D, Chen W, Wang J (2017) Light-induced variation in phenolic compounds in Cabernet Sauvignon grapes (Vitis vinifera L.) involves extensive transcriptome reprogramming of biosynthetic enzymes, transcription factors, and phytohormonal regulators. Front Plant Sci 8:547CrossRefGoogle Scholar
- Wang L, Long C-L (2003) Tissue culture of Boea hygrometrica. Plant Physiol Commun 39:233Google Scholar
- Xu W, Yang T, Dong X, Li D-Z, Liu A (2016) Genomic DNA methylation analyses reveal the distinct profiles in castor bean seeds with persistent endosperms. Plant Physiol 171:1242–1258Google Scholar