Xiong S, Chirala S S, Hsu M H, Wakil S J
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030, USA.
Proc Natl Acad Sci U S A. 1998 Oct 13;95(21):12260-5. doi: 10.1073/pnas.95.21.12260.
To investigate the regulation of the human fatty acid synthase gene by the thyroid hormone triiodothyronine, various constructs of the human fatty acid synthase promoter and the luciferase reporter gene were transfected in combination with plasmids expressing the thyroid hormone and the retinoid X receptors in HepG2 cells. The reporter gene was activated 25-fold by the thyroid hormone in the presence of the thyroid hormone receptor. When both the thyroid hormone and the retinoid X receptors were expressed in HepG2 cells, there was about a 100-fold increase in reporter gene expression. 5'-Deletion analysis disclosed two thyroid hormone response elements, TRE1 (nucleotides -870 to -650) and TRE2 (nucleotides -272 to -40), in the human fatty acid synthase promoter. The presence of thyroid hormone response elements in these two regions of the promoter was confirmed by cloning various fragments of these two regions in the minimal thymidine kinase promoter-luciferase reporter gene plasmid construct and determining reporter gene expression. The results of this cloning procedure and those of electrophoretic mobility shift assays indicated that the sequence GGGTTAcgtcCGGTCA (nucleotides -716 to -731) represents TRE1 and that the sequence GGGTCC (nucleotides -117 to -112) represents TRE2. The sequence of TRE1 is very similar to the consensus sequence of the thyroid hormone response element, whereas the sequence of TRE2 contains only a half-site of the thyroid hormone response element consensus motif because it lacks the direct repeat. The sequences on either side of TRE2 seem to influence its response to the thyroid hormone and retinoid X receptors.
为研究甲状腺激素三碘甲状腺原氨酸对人脂肪酸合酶基因的调控作用，将人脂肪酸合酶启动子的各种构建体与荧光素酶报告基因组合，转染至HepG2细胞中，并同时转染表达甲状腺激素受体和视黄酸X受体的质粒。在甲状腺激素受体存在的情况下，甲状腺激素可使报告基因的活性增强25倍。当HepG2细胞中同时表达甲状腺激素受体和视黄酸X受体时，报告基因的表达增加约100倍。5'-缺失分析揭示了人脂肪酸合酶启动子中有两个甲状腺激素反应元件，即TRE1（核苷酸-870至-650）和TRE2（核苷酸-272至-40）。通过将这两个区域的各种片段克隆到最小胸苷激酶启动子-荧光素酶报告基因质粒构建体中，并检测报告基因的表达，证实了启动子这两个区域中存在甲状腺激素反应元件。该克隆实验结果以及电泳迁移率变动分析结果表明，序列GGGTTAcgtcCGGTCA（核苷酸-716至-731）代表TRE1，序列GGGTCC（核苷酸-117至-112）代表TRE2。TRE1的序列与甲状腺激素反应元件的共有序列非常相似，而TRE2的序列仅包含甲状腺激素反应元件共有基序的半个位点，因为它缺乏直接重复序列。TRE2两侧的序列似乎影响其对甲状腺激素和视黄酸X受体的反应。
