Differential DNA methylation in regulation of deacetylvindoline-4-O-acetyl transferase ( DAT ) gene in Catharanthus rose
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ORIGINAL ARTICLE
Differential DNA methylation in regulation of deacetylvindoline-4-Oacetyl transferase (DAT) gene in Catharanthus roseus Nidarshana Chaturvedi Parashar1,4 • Gaurav Parashar2,4 • Harsh Nayyar3 • Rajat Sandhir1 Received: 6 February 2020 / Accepted: 20 August 2020 Ó Society for Plant Biochemistry and Biotechnology 2020
Abstract The present study investigates the role of DNA methylation in regulation of deacetylvindoline-4-O-acetyltransferase (DAT) gene in Madagascar periwinkle (Catharanthus roseus). Expression analysis showed significant upregulated DAT transcript (20 fold), demeter like (1.37 fold) and repressor of silencing-1 (1.47 fold) in leaf tissues of C. roseus as compared to root tissues. Methylases such as domain rearranged methyltransferase and chromomethyl transferase were downregulated suggesting involvement of DNA methylation in DAT regulation. DNA methylation signatures on DAT gene were further evaluated after treatment of leaves from C.roseus with 5-azacytidine (20 lM) for 5 and 10 days. Promoter DNA (- 11 to - 378 region) methylation analysis showed hypermethylation on two CHG and three CHH sites after day 10 of treatment. Qualitative methylation specific PCR of the coding region (? 153 to ? 322 region) showed hypomethylation after 10 days of 5-azacytidine treatment. Expression analysis showed upregulated DAT gene expression in the leaves 10 days post treatment, which correlated with changes observed in DNA methylation. Moreover, genomic 5-methylcytosine levels decreased from 0.65 to 0.33% after 10 days of 5-azacytidine treatment. However, no significant change was observed after 5 day of 5-azacytidine treatment as compared to control across different observational end points. The study clearly suggests that DAT gene expression may be regulated by differential DNA methylation in the promoter and coding regions. Further, hypomethylating agent such as 5-azacytidine holds potential to regulate the expression of DAT that regulates the terminal step in the indole alkaloid pathway leading to vindoline synthesis. Keywords Catharanthus roseus Vincristine Vinblastine DAT 5-azacytidine Alkaloids DNA methylation Abbreviations CMT3 Chromomethyl transferase 3 DAT Deacetylvindoline-4-O-acetyl transferase DML1 Demeter like 1 DRM2 Domain rearranged methyltransferase
MeJA ROS1 STR STS
& Rajat Sandhir [email protected]
Introduction
1
Department of Biochemistry, Panjab University, Basic Medical Sciences Block II, Sector 25, Chandigarh 160014, India
2
Laboratory of Epigenetics and Diseases, Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research, S.A.S. Nagar (Mohali), Punjab 160062, India
3
Department of Botany, Panjab University, Chandigarh 160014, India
4
Present Address: Department of Biotechnology, Maharishi Markandeshwar (Deemed to be University), Mullana, Ambala, Haryana 133207, India
Methyl jasmonate Repressor of silencing 1 Strictosidine synthase Stilbene synthase
Catharanthus roseus, commonly known as Madagascar periwinkle,
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