Berg Hakan, Modig Carina, Olsson Per-Erik
Department of Molecular Biology, Umea University, Umea, Sweden.
Reprod Biol Endocrinol. 2004 Sep 2;2:62. doi: 10.1186/1477-7827-2-62.
This study was performed to investigate stress effects on the synthesis of egg yolk precursor, vitellogenin (Vtg) in Arctic char (Salvelinus alpinus). In particular the effect of cortisol (F) was determined since this stress hormone has been suggested to interfere with vitellogenesis and is upregulated during sexual maturation in teleosts. Arctic char Vtg was purified and polyclonal antibodies were produced in order to develop tools to study regulation of vitellogenesis. The Vtg antibodies were used to develop an enzyme-linked immunosorbent assay. The corresponding Vtg cDNA was cloned from a hepatic cDNA library in order to obtain DNA probes to measure Vtg mRNA expression. Analysis of plasma from juvenile Arctic char, of both sexes, exposed to different steroids showed that production of Vtg was induced in a dose dependent fashion by 17beta-estradiol (E2), estrone and estriol. Apart from estrogens a high dose of F also upregulated Vtg. In addition, F, progesterone (P) and tamoxifen were tested to determine these compounds ability to modulate E2 induced Vtg synthesis at both the mRNA and protein level. Tamoxifen was found to inhibit E2 induced Vtg mRNA and protein upregulation. P did not alter the Vtg induction while F reduced the Vtg protein levels without affecting the Vtg mRNA levels. Furthermore the inhibition of Vtg protein was found to be dose dependent. Thus, the inhibitory effect of F on Vtg appears to be mediated at the post-transcriptional level.
本研究旨在调查应激对北极红点鲑(Salvelinus alpinus)卵黄前体卵黄蛋白原(Vtg)合成的影响。特别测定了皮质醇(F)的作用,因为这种应激激素被认为会干扰卵黄生成,并且在硬骨鱼的性成熟过程中会上调。纯化了北极红点鲑Vtg并制备了多克隆抗体,以开发研究卵黄生成调控的工具。利用Vtg抗体开发了一种酶联免疫吸附测定法。从肝脏cDNA文库中克隆了相应的Vtg cDNA,以获得用于测量Vtg mRNA表达的DNA探针。对暴露于不同类固醇的雌雄幼年北极红点鲑血浆的分析表明,17β-雌二醇(E2)、雌酮和雌三醇以剂量依赖性方式诱导Vtg的产生。除雌激素外,高剂量的F也上调了Vtg。此外,测试了F、孕酮(P)和他莫昔芬,以确定这些化合物在mRNA和蛋白质水平上调节E2诱导的Vtg合成的能力。发现他莫昔芬抑制E2诱导的Vtg mRNA和蛋白质上调。P没有改变Vtg的诱导,而F降低了Vtg蛋白水平,而不影响Vtg mRNA水平。此外,发现F对Vtg蛋白的抑制作用是剂量依赖性的。因此,F对Vtg的抑制作用似乎是在转录后水平介导的。