| Summary: | 博士 === 國立中興大學 === 生物科技學研究所 === 103 === In eukaryotes, the ubiquitin/26S proteasome system is one of the major mechanisms that act on the protein degradation. Proteins are tagged with ubiquitin as the signals to be degraded. In the cells, the ubiquitin-tagged proteins can be recognized and to regulate the life cycle of proteins. It was proposed that RING finger gene DAF-Like gene 1 (DAFL1) encodes an E3 protein ligase that plays a key role in 26S proteasome pathway in Arabidopsis thaliana. Based on phylogenetic analysis, DAFL1was closely related to DAF (DAD1-Activating Factor); however, the spatial specificities of DAFL1 and DAF gene expression patterns in Arabidopsis thaliana were different. In the early age of floral organs of Arabidopsis thaliana, DAFL1 specifically accumulated in the pistil and the ovule whereas DAF was expressed in stamen. It was suggested that DAFL1 and DAF might behave divergently in regulating plant development. Transgenic Arabidopsis plants that ectopically expressing DAFL1 RNAi and antisense caused indehiscent anthers and resulted in the unreleased pollen from the anthers. Similarly, ectopic expression of DAFL1-H135A caused indehiscent anthers in transgenic Arabidopsis plants. Pollens of wild-type Arabidopsis were crossed to the stigmas of sterile transgenic plants. At 7 days after pollination, the siliques of sterile transgenic plants developed and elongated. However, the amount of seeds of the transgenic plants is about a half of that of the wild-type Arabidopsis. Furthermore, transgenic plants that specifically expression of DAFL1-H135A in the pistil and the ovule produced dehiscent anthers, but the amount of seeds of the transgenic plants is about a half of that of the wild-type Arabidopsis. To discover the proteins that might interact with DAFL1, yeast-two hybrid system was performed to screen putative ones from flower buds cDNA library of Arabidopsis. One of the candidate genes, CAD1, was applied in the promoter assay. The result indicated that the gene expression patterns of CAD1 and DAFL1 was the same in ovule. In addition, transgenic plants that ectopic expressing CAD1 produced the amount of seeds is about a half of that of the wild-type Arabidopsis. CAD1 is a key enzyme in the process of lignin synthesis and has been thought to be involved in the lignification of siliques. Interestingly, DAFL1 and CAD1 also highly accumulated in the non-lignification site such as in the carpel. Our results revealed that DAFL1 degrades CAD1 and inhibits lignin biosynthesis at the early stage of pistil development.
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