Developmental- and tissue-specific expression of NbCMT3-2 encoding a chromomethylase in Nicotiana benthamiana

• Main Authors: Yu-Ting Lin, 林郁婷
• Other Authors: Shih-Feng Fu
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/08919418895148013477

碩士 === 國立彰化師範大學 === 生物學系 === 103 === DNA methylation is a heritable epigenetic process controlling gene expression and developmental programs in various organisms. The chromomethylase (CMT) protein family is unique to plants and controls non-CpG methylation. Here, we investigated the developmental expression of CMT3-2 in Nicotiana benthamiana (NbCMT3-2) and its significance by analyzing plants with silencing of NbCMT3-2 and analyzing leaf tissues transiently expressing the N-terminal polypeptide. Alignment of the NbCMT3-2 amino acid sequence with other plant CMT3s showed a specific N-terminal extension required for nuclear localization. Transient expression of the N-terminal polypeptide in N. benthamiana resulted in the formation of chlorotic lesions, which indicates its vital role in leaf development. NbCMT3-2 was expressed mainly in proliferating tissues such as the shoot apex and developing leaves. We generated transgenic N. benthamiana harboring a fusion reporter construct linking the NbCMT3-2 promoter region and -glucuronidase (GUS) reporter (pNbCMT3-2::GUS) to analyze the tissue-specific expression of NbCMT3-2. NbCMT3-2 was expressed in shoot and root apical meristem and leaf primordia in young seedlings and highly expressed in developing leaves and ovary as well as lateral buds in mature plants. Virus-induced gene silencing used to knock down NbCMT3-2 expression led to partial loss of genomic CHG DNA methylation. Silencing NbCMT3-2 interfered with leaf development and the expression of genes involved in jasmonate homeostasis. The differential roles between NbCMT3 and NbCMT3-2 were investigated and compared. We reveal the expression patterns of NbCMT3-2 in proliferating tissues. NbCMT3-2 may play an essential role in leaf development by modulating jasmonate pathways.