Molecular cloning of genes for sex reversal in amphibians

分离编码蛙、Rana rugosa calnexin（2.2 kb）和calreticulin（2.4 kb）的cDNA克隆并测序。CLX基因仅在肝脏中弱表达，而CLT基因在成年青蛙的所有组织中均表达。年轻的青蛙在阶段XXV产生了一个非常强烈的信号，CLT基因表达的卵巢，但不是在睾丸。丙酸睾酮（TP）诱导蝌蚪雄性化时，CLT基因在性腺中的表达受到显著抑制。在免疫组织学研究中，CLT在18-19期蝌蚪的卵母细胞中未检测到，但在变态后的青蛙卵母细胞的整个细胞质中检测到。随着卵黄颗粒的积累，这种蛋白质被挤压向细胞核。我们的研究结果表明，CLT，而不是CLX基因在卵母细胞中表达的青蛙，R. rugosa成熟过程中。这种青蛙的雄性化至少部分与分化性腺中CLT基因表达的抑制有关。此外，还分离并测序了一个2.2kb的Ad 4 BP cDNA，该基因是已知的分化性腺和肾上腺的关键基因。Ad 4 BP基因在青蛙发育过程中的表达及其基因表达的性激素调控正在研究中。

cDNA clones encoding the frog, Rana rugosa calnexin (2.2 kb) and calreticulin (2.4 kb) were isolated and sequenced. The CLX gene was weakly expressed only in the liver, while the CLT gene was expressed in all the tissues examined of adult frogs. Young frogs at stage XXV produced an extremely strong signal of the CLT gene expression in the ovary, but not in the testis. When the masculinization was induced by testosterone propionate (TP) , the CLT gene expression was significantly inhibited in gonads of tadpoles. In immunohistological studies, CLT was not appreciably detected in oocytes in ggonads of tadpoles at stage 18-19, but detected in the whole cytoplasm of growing oocytes of frogs after metamorphosis. This protein then was squeezed toward the nucleus as yolk granules were accumulated. Our results suggest that the CLT,but not CLX gene is expressed in oocytes during maturation in the frog, R.rugosa. The masculinization of this frog is at least partly associated with the suppression of the CLT gene expression in differentiating gonads. In addition, A 2.2-kb cDNA of Ad4BP,which is known to be a key gene for differentiation of gonad and adrenal in mammalians, was also isolated and sequenced. The Ad4BP gene expression during development of frogs and a sex hormonal regulation of its gene expression are now under investigations.