Averyhart-Fullard V, Jaffe R C
Department of Physiology and Biophysics, University of Illinois, Chicago 60680.
Mol Endocrinol. 1990 Oct;4(10):1556-63. doi: 10.1210/mend-4-10-1556.
Thyroid hormones are responsible for the specific biochemical and structural changes that occur during amphibian metamorphosis. In this study we screened a series of cDNAs from a library constructed from T4-treated premetamorphic tadpole liver poly(A)+ RNA in order to identify a clone that could be used to study the influence of T3 on liver-specific gene expression during Rana catesbeiana metamorphosis. The cDNA from one clone exhibited a greater degree of hybridization to liver RNA from thyroid hormone-treated tadpoles than untreated tadpoles and no hybridization to RNA from tail fins of tadpoles of either group. On Northern blots, the mRNA to which the cDNA hybridized was 2.3 kilobases in size. The pattern of hybridization to genomic DNA digested by various restriction enzymes was consistent with the presence of a single gene. Using slot blot analysis we found that the mRNA levels first rose above basal levels only after 5 days of immersion of tadpoles in 12.5 micrograms/liter T3. The mRNA levels increased approximately 10-fold after 7 and 9 days of treatment. Frog livers had mRNA levels that were intermediate between those in untreated tadpoles and tadpoles immersed in T3 for 7 days. Sequence analysis revealed a significant degree of homology to serum albumin and alpha-fetoprotein. While it is known that serum albumin levels rise dramatically during metamorphosis in Rana species, presumably playing a critical role in maintaining water and electrolyte balance during the animals' terrestrial phase, the molecular basis of the induction has not been fully explained.
甲状腺激素负责两栖动物变态过程中发生的特定生化和结构变化。在本研究中,我们从用T4处理的变态前蝌蚪肝脏poly(A)+ RNA构建的文库中筛选了一系列cDNA,以鉴定一个可用于研究T3对牛蛙变态过程中肝脏特异性基因表达影响的克隆。来自一个克隆的cDNA与甲状腺激素处理过的蝌蚪的肝脏RNA的杂交程度高于未处理的蝌蚪,且与两组蝌蚪尾鳍的RNA均无杂交。在Northern印迹上,与该cDNA杂交的mRNA大小为2.3千碱基。与各种限制酶消化的基因组DNA的杂交模式与单个基因的存在一致。使用狭缝印迹分析,我们发现只有在蝌蚪浸泡在12.5微克/升T3中5天后,mRNA水平才首次升至基础水平之上。处理7天和9天后,mRNA水平增加了约10倍。青蛙肝脏的mRNA水平介于未处理的蝌蚪和浸泡在T3中7天的蝌蚪之间。序列分析显示与血清白蛋白和甲胎蛋白有高度同源性。虽然已知在蛙类变态过程中血清白蛋白水平会急剧上升,推测在动物陆地阶段维持水和电解质平衡中起关键作用,但诱导的分子基础尚未得到充分解释。