The ability for plant regeneration from dedifferentiated cells opens up the possibility for molecular bioengineering to produce crops with desirable traits. Developmental and environmental signals that control cell totipotency are regulated by gene expression via dynamic chromatin remodeling. Using a mass spectrometry-based approach, we investigated epigenetic changes to the histone proteins during callus formation from roots and shoots of Arabidopsis thaliana seedlings. Increased levels of the histone H3.3 variant were found to be the major and most prominent feature of 20-day calli, associated with chromatin relaxation. The methylation status in root- and shoot-derived calli reached the same level during long-term propagation, whereas differences in acetylation levels provided a long-lasting imprint of root and shoot origin. On the other hand, epigenetic signs of origin completely disappeared during 20 days of calli propagation in the presence of histone deacetylase inhibitors (HDACi), sodium butyrate, and trichostatin A. Each HDACi affected the state of post-translational histone modifications in a specific manner; NaB-treated calli were epigenetically more similar to root-derived calli, and TSA-treated calli resembled shoot-derived calli.

Mendel Centre for Plant Genomics and Proteomics Central European Institute of Technology Masaryk University 62500 Brno Czech Republic

National Centre for Biomolecular Research Faculty of Science Masaryk University 62500 Brno Czech Republic

PSI spol s r o 66424 Drásov Czech Republic

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Long Y., Yang Y., Pan G., Shen Y. New Insights Into Tissue Culture Plant-Regeneration Mechanisms. Front. Plant Sci. 2022;13:926752. doi: 10.3389/fpls.2022.926752. PubMed DOI PMC

Skoog F., Miller C.O. Chemical regulation of growth and organ formation in plant tissues cultured in vitro. Symp. Soc. Exp. Biol. 1957;11:118–130. PubMed

Lardon R., Geelen D. Natural Variation in Plant Pluripotency and Regeneration. Plants. 2020;9:1261. doi: 10.3390/plants9101261. PubMed DOI PMC

Benkova E., Bielach A. Lateral root organogenesis—From cell to organ. Curr. Opin. Plant Biol. 2010;13:677–683. doi: 10.1016/j.pbi.2010.09.006. PubMed DOI

Che P., Lall S., Howell S.H. Developmental steps in acquiring competence for shoot development in Arabidopsis tissue culture. Planta. 2007;226:1183–1194. doi: 10.1007/s00425-007-0565-4. PubMed DOI

Sugimoto K., Jiao Y., Meyerowitz E.M. Arabidopsis regeneration from multiple tissues occurs via a root development pathway. Dev. Cell. 2010;18:463–471. doi: 10.1016/j.devcel.2010.02.004. PubMed DOI

Lee K., Park O.S., Seo P.J. RNA-Seq Analysis of the Arabidopsis Transcriptome in Pluripotent Calli. Mol. Cells. 2016;39:484–494. doi: 10.14348/molcells.2016.0049. PubMed DOI PMC

Klemm S.L., Shipony Z., Greenleaf W.J. Chromatin accessibility and the regulatory epigenome. Nat. Rev. Genet. 2019;20:207–220. doi: 10.1038/s41576-018-0089-8. PubMed DOI

Pfluger J., Wagner D. Histone modifications and dynamic regulation of genome accessibility in plants. Curr. Opin. Plant Biol. 2007;10:645–652. doi: 10.1016/j.pbi.2007.07.013. PubMed DOI PMC

Liu C., Lu F., Cui X., Cao X. Histone methylation in higher plants. Annu. Rev. Plant Biol. 2010;61:395–420. doi: 10.1146/annurev.arplant.043008.091939. PubMed DOI

Feher A. Somatic embryogenesis—Stress-induced remodeling of plant cell fate. Biochim. Biophys. Acta. 2015;1849:385–402. doi: 10.1016/j.bbagrm.2014.07.005. PubMed DOI

Ikeuchi M., Iwase A., Rymen B., Harashima H., Shibata M., Ohnuma M., Breuer C., Morao A.K., de Lucas M., De Veylder L., et al. PRC2 represses dedifferentiation of mature somatic cells in Arabidopsis. Nat. Plants. 2015;1:15089. doi: 10.1038/nplants.2015.89. PubMed DOI

Mozgova I., Munoz-Viana R., Hennig L. PRC2 Represses Hormone-Induced Somatic Embryogenesis in Vegetative Tissue of Arabidopsis thaliana. PLoS Genet. 2017;13:e1006562. doi: 10.1371/journal.pgen.1006562. PubMed DOI PMC

He C., Chen X., Huang H., Xu L. Reprogramming of H3K27me3 is critical for acquisition of pluripotency from cultured Arabidopsis tissues. PLoS Genet. 2012;8:e1002911. doi: 10.1371/journal.pgen.1002911. PubMed DOI PMC

Bannister A.J., Kouzarides T. Regulation of chromatin by histone modifications. Cell Res. 2011;21:381–395. doi: 10.1038/cr.2011.22. PubMed DOI PMC

Lee K., Park O.S., Jung S.J., Seo P.J. Histone deacetylation-mediated cellular dedifferentiation in Arabidopsis. J. Plant Physiol. 2016;191:95–100. doi: 10.1016/j.jplph.2015.12.006. PubMed DOI

Kim J.Y., Yang W., Forner J., Lohmann J.U., Noh B., Noh Y.S. Epigenetic reprogramming by histone acetyltransferase HAG1/AtGCN5 is required for pluripotency acquisition in Arabidopsis. EMBO J. 2018;37:e98726. doi: 10.15252/embj.201798726. PubMed DOI PMC

Mottamal M., Zheng S., Huang T.L., Wang G. Histone deacetylase inhibitors in clinical studies as templates for new anticancer agents. Molecules. 2015;20:3898–3941. doi: 10.3390/molecules20033898. PubMed DOI PMC

Earley K.W., Shook M.S., Brower-Toland B., Hicks L., Pikaard C.S. In vitro specificities of Arabidopsis co-activator histone acetyltransferases: Implications for histone hyperacetylation in gene activation. Plant J. 2007;52:615–626. doi: 10.1111/j.1365-313X.2007.03264.x. PubMed DOI

Chen Z.J., Pikaard C.S. Epigenetic silencing of RNA polymerase I transcription: A role for DNA methylation and histone modification in nucleolar dominance. Genes Dev. 1997;11:2124–2136. doi: 10.1101/gad.11.16.2124. PubMed DOI PMC

Pagano A., de Sousa Araujo S., Macovei A., Dondi D., Lazzaroni S., Balestrazzi A. Metabolic and gene expression hallmarks of seed germination uncovered by sodium butyrate in Medicago truncatula. Plant Cell Environ. 2019;42:259–269. doi: 10.1111/pce.13342. PubMed DOI

Tanaka M., Kikuchi A., Kamada H. The Arabidopsis histone deacetylases HDA6 and HDA19 contribute to the repression of embryonic properties after germination. Plant Physiol. 2008;146:149–161. doi: 10.1104/pp.107.111674. PubMed DOI PMC

Ledvinova D., Mikulasek K., Kucharikova H., Brabencova S., Fojtova M., Zdrahal Z., Lochmanova G. Filter-Aided Sample Preparation Procedure for Mass Spectrometric Analysis of Plant Histones. Front. Plant Sci. 2018;9:1373. doi: 10.3389/fpls.2018.01373. PubMed DOI PMC

Lochmanova G., Ihnatova I., Kucharikova H., Brabencova S., Zachova D., Fajkus J., Zdrahal Z., Fojtova M. Different Modes of Action of Genetic and Chemical Downregulation of Histone Deacetylases with Respect to Plant Development and Histone Modifications. Int. J. Mol. Sci. 2019;20:5093. doi: 10.3390/ijms20205093. PubMed DOI PMC

Jacob Y., Bergamin E., Donoghue M.T.A., Mongeon V., LeBlanc C., Voigt P., Underwood C.J., Brunzelle J.S., Michaels S.D., Reinberg D., et al. Selective Methylation of Histone H3 Variant H3.1 Regulates Heterochromatin Replication. Science. 2014;343:1249–1253. doi: 10.1126/science.1248357. PubMed DOI PMC

Johnson L., Mollah S., Garcia B.A., Muratore T.L., Shabanowitz J., Hunt D.F., Jacobsen S.E. Mass spectrometry analysis of histone H3 reveals distinct combinations of post-translational modifications. Nucleic Acids Res. 2004;32:6511–6518. doi: 10.1093/nar/gkh992. PubMed DOI PMC

Bednarek P.T., Orlowska R. Plant tissue culture environment as a switch-key of (epi)genetic changes. Plant Cell Tiss. Org. 2020;140:245–257. doi: 10.1007/s11240-019-01724-1. DOI

Miguel C., Marum L. An epigenetic view of plant cells cultured: Somaclonal variation and beyond. J. Exp. Bot. 2011;62:3713–3725. doi: 10.1093/jxb/err155. PubMed DOI

Us-Camas R., Rivera-Solís G., Duarte-Aké F., De-la-Peña C. In vitro culture: An epigenetic challenge for plants. Plant Cell Tiss. Org. 2014;118:187–201. doi: 10.1007/s11240-014-0482-8. DOI

Birnbaum K.D., Roudier F. Epigenetic memory and cell fate reprogramming in plants. Regeneration. 2017;4:15–20. doi: 10.1002/reg2.73. PubMed DOI PMC

Hemenway E.A., Gehring M. Epigenetic Regulation During Plant Development and the Capacity for Epigenetic Memory. Annu. Rev. Plant Biol. 2023;74:87–109. doi: 10.1146/annurev-arplant-070122-025047. PubMed DOI PMC

Xu K., Liu J., Fan M., Xin W., Hu Y., Xu C. A genome-wide transcriptome profiling reveals the early molecular events during callus initiation in Arabidopsis multiple organs. Genomics. 2012;100:116–124. doi: 10.1016/j.ygeno.2012.05.013. PubMed DOI

Ahmad K., Henikoff S. The histone variant H3.3 marks active chromatin by replication-independent nucleosome assembly. Mol. Cell. 2002;9:1191–1200. doi: 10.1016/S1097-2765(02)00542-7. PubMed DOI

Nie X., Wang H., Li J., Holec S., Berger F. The HIRA complex that deposits the histone H3.3 is conserved in Arabidopsis and facilitates transcriptional dynamics. Biol. Open. 2014;3:794–802. doi: 10.1242/bio.20148680. PubMed DOI PMC

Stroud H., Otero S., Desvoyes B., Ramírez-Parra E., Jacobsen S.E., Gutierrez C. Genome-wide analysis of histone H3.1 and H3.3 variants in Arabidopsis thaliana. Proc. Natl. Acad. Sci. USA. 2012;109:5370–5375. doi: 10.1073/pnas.1203145109. PubMed DOI PMC

Zhang H., Guo F., Qi P., Huang Y., Xie Y., Xu L., Han N., Bian H. OsHDA710-Mediated Histone Deacetylation Regulates Callus Formation of Rice Mature Embryo. Plant Cell Physiol. 2020;61:1646–1660. doi: 10.1093/pcp/pcaa086. PubMed DOI

Bie X.M., Dong L., Li X.H., Wang H., Gao X.Q., Li X.G. Trichostatin A and sodium butyrate promotes plant regeneration in common wheat. Plant Signal. Behav. 2020;15:1820681. doi: 10.1080/15592324.2020.1820681. PubMed DOI PMC

Furuta K., Kubo M., Sano K., Demura T., Fukuda H., Liu Y.G., Shibata D., Kakimoto T. The CKH2/PKL chromatin remodeling factor negatively regulates cytokinin responses in Arabidopsis calli. Plant Cell Physiol. 2011;52:618–628. doi: 10.1093/pcp/pcr022. PubMed DOI

Choi S.H., Ahn W.S., Lee M.H., Jin D.M., Lee A., Jie E.Y., Ju S.J., Ahn S.J., Kim S.W. Effects of TSA, NaB, Aza in Lactuca sativa L. protoplasts and effect of TSA in Nicotiana benthamiana protoplasts on cell division and callus formation. PLoS ONE. 2023;18:e0279627. doi: 10.1371/journal.pone.0279627. PubMed DOI PMC

Kolarova H., Vitecek J., Cerna A., Cernik M., Pribyl J., Skladal P., Potesil D., Ihnatova I., Zdrahal Z., Hampl A., et al. Myeloperoxidase mediated alteration of endothelial function is dependent on its cationic charge. Free Radic. Biol. Med. 2021;162:14–26. doi: 10.1016/j.freeradbiomed.2020.11.008. PubMed DOI

Kucharikova H., Plskova Z., Zdrahal Z., Fojtova M., Kerchev P., Lochmanova G. Quantitative Analysis of Posttranslational Modifications of Plant Histones. Methods Mol. Biol. 2022;2526:241–257. doi: 10.1007/978-1-0716-2469-2_18. PubMed DOI

Aitchison J. The Statistical-Analysis of Compositional Data. J. R. Stat. Soc. B. 1982;44:139–177. doi: 10.1111/j.2517-6161.1982.tb01195.x. DOI

Perez-Riverol Y., Bai J., Bandka C., Hewapathirana S., García-Seisdedos D., Kamatchinathan S., Kundu D., Prakash A., Frericks-Zipper A., Eisenacher M., et al. The PRIDE database resources in 2022: A Hub for mass spectrometry-based proteomics evidences. Nucleic Acids Res. 2022;50:D543–D552. doi: 10.1093/nar/gkab1038. PubMed DOI PMC

Arg-C Ultra Simplifies Histone Preparation for LC-MS/MS