碩士 / 國立臺灣海洋大學 / 生物科技研究所 / 93 / Abstract
Krüppel-like factors (KLFs) encoding transcription factors have been found in many organisms. They contain three highly conserved tandem zinc finger DNA binding motifs in the C-terminal region and unique functional domain in the N-terminal coding region. Studies indicated that KLFs are multi-functional protein and play key roles in cell proliferation, differentiation, as well as regulation of cell cycle and repression of tumor. Recently, zebrafish becomes a model organism for studying vertebrate embryogenesis. However, derailed molecular mechanisms regarding morphogenesis of digestive tract development are still lacking. Therefore, in this thesis we investigated possible roles of zebrafish KLF in digestive tract development during embryogenesis.
We have previously cloned a zebrafish KLF4a cDNA, which contains 2119bp and encodes 396 amino acids. Amino acid sequence comparison showed that zebrafish KLF4a shares 65.8％ sequence similarity with mouse KLF4. Zebrafish KLF4a also contains acidic activation domain in the N-terminus and nuclear localization signal and three highly conserved zinc finger DNA binding domains in the C-terminal region. Phylogenetic tree analysis grouped zebrafish KLF4a with human and mouse KLF4 in the same branch. Whole mount in situ hybridization revealed KLF4a mRNA first detected in 1 cell stage and was expressed in blastula period. KLF4a was expressed in the dorsal shield and axial mesendoderm during gastrula. During segmentation period, KLF4a was expressed in the otic primordial, brain and somite. After 24 hpf KLF4a was expressed in the eyes, ventral region of brain, midbrain-hindbrain boundary (MHB), ventricular zone, otic vesicle, somites, pharynx, liver, exocrine pancreas and gut. Frozen sectioning of 96 hpf embryos hybridized with KLF4a RNA probe showed KLF4a was mainly localized in epithelial cell of gut.
Using KLF4a antisense morpholino oligomer to inhibit the function of KLF4a, development of MHB and dorsal-ventral axis of brain in morphant were affected in embryos form 24 hpf to 72 hpf. In the meantime, the development of eyes and semi-circular canal was delayed. After 72 hpf, cavity of foregut, folding of gut and development of swim-bladder were affected. Conparison of expression pattern of different digestive tract marker genes in KLF4a morphant and wild-type embryos showed that the expression level of dlx7 in esophagus and insulin in endocrine pancreas were not alterd. However, the expression level and domain of IFABP in the liver and foregut, LFABP in the liver and trypsin in the exocrine pancreas were decreased. On the contrary, expression of shh in the month and pharynx were inhibited by KLF4a. Overall, our results showed that KLF4a is homologue of mammalian KLF4 and it plays an important role in the development of liver, exocrine pancreas and gut.
| Identifer | oai:union.ndltd.org:TW/093NTOU5111019 |
| Date | January 2005 |
| Creators | Guo-Bin Li, 李國斌 |
| Contributors | Chin-Hwa Hu, Sheng-Ping L. Hwang, 胡清華, 黃聲蘋 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 111 |
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