Functional characterization of Nicotiana benthamiana chromomethylase NbCMT3-2 gene in defense to ToLCTWV

• Main Authors: Wei,Jyuan-Yu, 魏雋瑀
• Other Authors: Fu, Shih-Feng
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/84pu45

碩士 === 國立彰化師範大學 === 生物學系 === 105 === Plants are constantly exposed to variety of the biotic and abiotic stresses throughout their life cycle. Epigenetic systems including histone modifications and DNA methylation are crucial for maintaining chromatin structure and genome reprogramming in response to environmental stresses. DNA methylation is a heritable epigenetic process controlling gene expression and developmental programs in various organisms. Methylation of cytosine residues in DNA is enzymatically catalyzed by DNA methyltransferases (DNMT). Plants possess 3 types of DNMTs that differ in protein structures and functions (Methyltransferases, MET; Chromomethylase, CMT3; Domains rearranged methyltransferases, DRM). However, the roles of plant DNMTs in defense against DNA virus such as geminivirus remain to be elusive. In this study, we generated the NbCMT3-2 knockdown and overexpressing lines by virus‐induced gene silencing (VIGS) and by overexpression of the target gene, respectively. The NbCMT3-2–silencing Nicotiana benthamiana plants (VIGS-NbCMT3-2 transient lines) were more susceptible to infection with tomato leaf curl Taiwan virus (ToLCTWV) than the control plants. Expression of the gene encoding the coat protein (CP) of ToLCTWV was enhanced in NbCMT3-2–silenced plants in comparison with control plants. NbCMT3-2–silenced plants exhibited susceptibility to ToLCTWV infection that correlates with reduced cytosine methylation of the viral CP gene promoter. The results indicate that N. benthamiana plants with decrease in NbCMT3-2 expression was more susceptible to ToLCTWV infection. In contrast, resistance to ToLCTWV was enhanced in transgenic N. benthamiana plants overexpressing NbCMT3-2 (OX-NbCMT3-2 and HA-NbCMT3-2 stable transgenic lines). The NbCMT3-2 overexpressing–plants exhibited mild symptoms when infected with ToLCTWV. Taken together, this study demonstrated the vital roles of CMT3-2–mediated DNA methylation in resistance to geminiviruses.