Immunohistochemical detection and biological activities of CYP17 (P450c17) in the indifferent gonad of the frog Rana rugosa
Immunohistochemical detection and biological activities of CYP17 (P450c17) in the indifferent gonad of the frog Rana rugosa
复制标题
DOI：
10.1016/j.jsbmb.2008.07.002
复制
发表时间：
2008-11-01
影响因子：
4.1
通讯作者：
Nakamura, Masahisa
中科院分区：
文献类型：
--
作者：
Sakurai, Nana;Maruo, Koichi;Nakamura, Masahisa
性类固醇在各种脊椎动物的性腺分化中起着至关重要的作用。然而，对于两栖动物性腺性别分化过程中类固醇代谢酶的定位和生物活性知之甚少。在本研究中，我们通过实时RT-PCR分析发现，在雌雄雌雄蝌蚪的性腺分化过程中，雄性蝌蚪的不同性腺中关键甾体生成酶之一CYP17的表达高于雌性蝌蚪，而雄性和雌性蝌蚪性腺中3 β HSD mRNA的表达水平没有差异。接下来，我们分别使用三种针对CYP77、3 β HSD和17 β HSD的特异性抗体检测了CYP17、3 β HSD和17 β HSD在不同性腺和分化性腺中的定位。CYP17、3 β HSD和17 β HSD在雄性性腺的体细胞和睾丸间质细胞中表达阳性。在性腺分化过程中也检测了CYP17的酶活性。[H-3]孕酮（Prog）在雄性和雌性的不同性腺中转化为[H-3]雄烯二酮（AE），但其转化率在雄性中高于雌性。此外，荧光原位杂交（FISH）分析显示，CYP77基因位于第9染色体的q臂上，表明CYP17是常染色体。综上所述，CYP17蛋白在褐毛鼠性腺分化过程中由不同性腺的体细胞合成，并且在雄性褐毛鼠中将Prog转化为AE的过程中比雌性褐毛鼠更为活跃。这些数据表明CYP77可能参与了该物种性别分化过程中睾丸的形成。(C) 2008 Elsevier Ltd版权所有。
Sex steroids play a crucial role in the gonad differentiation in various species of vertebrates. However, little is known regarding the localization and biological activity of steroid-metabolizing enzymes during gonadal sex differentiation in amphibians. In the present study, we showed by real-time RT-PCR analysis that the expression of CYP17, one of the key steroidogenic enzymes, was higher in the indifferent gonad during sex differentiation in male than in female tadpoles of Rana rugosa but that there was no difference detected in the 3 beta HSD mRNA level between the male and female gonads. We next examined the localization of CYP17, 3 beta HSD and 17 beta HSD in the indifferent and differentiating gonads by using three kinds of antibodies specific for CYP77, 3 beta HSD and 17 beta HSD, respectively. Positive signals for CYP17, 3 beta HSD and 17 beta HSD were observed in somatic cells of the indifferent gonad of males and in the interstitial cell of the testis. The enzymatic activity of CYP17 was also examined in the gonad during sex differentiation in this species. [H-3]Progesterone (Prog) was converted to [H-3]androstenedione (AE) in the indifferent gonad in males and females, but the rate of its conversion was higher in males than in females. Moreover, fluorescence in situ hybridization (FISH) analysis revealed that the CYP77 gene was located on the q arm of chromosome 9, indicating that CYP17 was autosomal in R. rugosa. Taken together, the results demonstrate that the CYP17 protein is synthesized in somatic cells of the indifferent gonad during gonadal sex differentiation in R. rugosa and that it is more active in converting Prog to AE in males than in females. The data suggest that CYP77 may be involved in testicular formation during sex differentiation in this species. (C) 2008 Elsevier Ltd. All rights reserved.