Characterization and Functional Analysis of NAC-like and ERD Genes in Arabidopsis thaliana

• Main Authors: Chiu-Sien Ng, 吳秋嫺
• Other Authors: 楊長賢
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/06020839669141106691

碩士 === 中興大學 === 生物科技學研究所 === 99 === NAC (NAM, ATAF1,2 and CUC2 ) proteins have been considered as one of the plant-specific transcription factors that are involved in the regulation of diverse plant developmental processes. Two Arabidopsis NAC-like genes AtNACL2 and AtNACL10 were isolated and analyzed in this study. AtNACL2 and AtNACL10 were highly expressed in the leaves and flowers. Interestingly, AtNACL10 is also highly detected in root and inflorescence in Arabidopsis thaliana. T-DNA mutants of AtNACL2 and AtNACL10 were phenotypically indistinguishable from wild-type. To solve the problem caused by functional redundancy, the chimeric repressor silencing technology (CRES-T) was used. In 35S::AtNACL2-SRDX plants, the abnormal phenotype with serrated leaves was observed. 35S::AtNACL10-SRDX plants exhibited dramatic leaf curvature and multiple branches with secondary inflorescence or flowers at the same position of an internode. In addition, 35S::AtNACL2-SRDX and 35S::AtNACL10-SRDX showed serious sterility due to the production of short stamen and indehiscence of the anther. Further analysis indicated that these mutant phenotypes were correlated with the alteration of the expression for genes involved in meristematic development such as FAS (FASCIATA), WUS (WUSCHEL) and leaf development such as KNAT1(Knotted-like from Arabidopsis thaliana1), IAMT1(IAA carboxyl methyltransferase1). The results suggest that AtNACL2 and AtNACL10 are key regulators in regulating leaf morphology and floral development in Arabidopsis thaliana. A conserved sequence of CX2CX3FX5LX2HX3H identified in the Cys2/His2 zinc finger region of the AtEPF1 was present in EPF family genes. Ectopic expression of AtEPF1 in transgenic Arabidopsis plants showed phenotypes by significantly reducing the plant size and producing curled and dehydrated leaves. To investigate the target genes regulated by AtEPF1, microarray analysis was performed. Two of AtEPF1 regulated genes, ERD4 (EARLY-RESPONSIVE TO DEHYDRATION 4) and ERD7, were further analyzed. ERD4 and ERD7 were originally described as a rapidly drought-responsive gene in Arabidopsis. When exposing plants to progressive drought stress, we obtained elevated levels of the ERD4 and ERD7. In ERD4::GUS plants, GUS activity was detected in cotyledons, roots, hydathodes, sepal, style and pollens. In ERD7::GUS plants, the GUS activity was detected in cotyledons, roots, style and pollens. Interestingly, GUS activity in ERD4::GUS plants was detected in leaf veins by drought treatment. Ectopic expression of ERD7 showed novel phenotypes by reducing the plant size and producing yellow wrinkled leaf. Further analysis indicated that the chlorophyll relative contents also decreased, lipid peroxidation tended to increase and ERF1 (ETHYLENE-RESPONSE-FACTOR 1) expression up-regulated. The present evidence indicated that ERD4 and ERD7 may play roles in protection of the plants with water retention. These results support our speculation for the ERD4 and ERD7 function based on its regulation by AtEPF1.