Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Science, Southwest University, Chongqing, China.
Biol Reprod. 2012 May 31;86(5):163, 1-10. doi: 10.1095/biolreprod.111.096248. Print 2012 May.
Insulin-like growth factors (Igfs) are implicated in a wide variety of physiological roles in teleost gonadal development and reproduction. In the present study, igf3 mRNA expression in the tilapia ovary was found to be higher than in the testis from 5 to 40 days after hatching (dah) but was lower than that in testis from 50 to 70 dah. Consistently, Igf3 protein signal was detected in the somatic cells of XX and XY gonads from 10 dah until adulthood by immunohistochemistry, using a specific Igf3 polyclonal antibody. Incubation of ovarian and testicular cells in primary culture with recombinant Igf3 significantly increased nr5a1, foxl2, dmrt1, cyp19a1a, cyp11a1, cyp11b2, hsd3b2 , and cyp17a1 expression in a time- and dose-dependent manner. Promoter analysis using luciferase assays in HEK293 cells revealed that igf3 promoter activity was directly activated by Nr5a1 (Sf1) and further enhanced by Foxl2, Nr0b1a (Dax1), and Nr0b1b (Dax2) but repressed by Dmrt1 and estrogen receptor (Esr1, Esr2a, or Esr2b) along with 17beta-estradiol treatment. In addition, igf3 promoter activity was increased slightly by forskolin treatment alone but synergistically up-regulated by transfection with nr5a1. These in vitro results correlated well with the expression profile of igf3 during early gonad differentiation. Our results indicated that igf3 is involved in fish gonad steroidogenesis because of its ability to regulate the expression of foxl2, dmrt1, and nr5a1 and steroidogenic enzymes. The expression of igf3 is in turn regulated by transcription factors Foxl2, Dmrt1, and Nr5a1, as well as by 17beta-estradiol treatment.
胰岛素样生长因子（Igfs）在鱼类性腺发育和生殖的各种生理作用中都有涉及。在本研究中，发现从孵化后 5 天到 40 天（dah），罗非鱼卵巢中的 igf3 mRNA 表达高于睾丸，但低于 50 至 70 dah 的睾丸。同样，通过免疫组织化学，使用特异性 igf3 多克隆抗体，在 10 dah 至成年期的 XX 和 XY 性腺的体细胞中检测到 Igf3 蛋白信号。卵巢和睾丸细胞在原代培养中与重组 Igf3 孵育，以时间和剂量依赖的方式显著增加 nr5a1、foxl2、dmrt1、cyp19a1a、cyp11a1、cyp11b2、hsd3b2 和 cyp17a1 的表达。在 HEK293 细胞中使用荧光素酶测定进行的启动子分析表明，igf3 启动子活性直接被 Nr5a1（Sf1）激活，并进一步被 Foxl2、Nr0b1a（Dax1）和 Nr0b1b（Dax2）增强，但被 Dmrt1 和雌激素受体（Esr1、Esr2a 或 Esr2b）以及 17β-雌二醇处理抑制。此外，igf3 启动子活性单独用 forskolin 处理略有增加，但与 Nr5a1 转染协同上调。这些体外结果与早期性腺分化过程中 igf3 的表达谱很好地相关。我们的结果表明，igf3 参与鱼类性腺类固醇生成，因为它能够调节 foxl2、dmrt1 和 nr5a1 和类固醇生成酶的表达。igf3 的表达反过来又受到转录因子 Foxl2、Dmrt1 和 Nr5a1 的调节，以及 17β-雌二醇处理的调节。
