Hiruma K, Böcking D, Lafont R, Riddiford L M
Department of Zoology, University of Washington, Seattle 98195-1800, USA.
Gen Comp Endocrinol. 1997 Jul;107(1):84-97. doi: 10.1006/gcen.1997.6901.
To determine which ecdysteroids may be biologically active in the larval epidermis of the tobacco hornworm, Manduca sexta, we studied the action of several known ecdysteroids and metabolites on the expression of the genes encoding the ecdysone receptor (EcR), Manduca hormone receptor 3 (MHR3), dopa decarboxylase (DDC), and a larval cuticle protein (LCP-14). Both Day 2 fourth- and Day 2 fifth-instar larval epidermis contained significant 3 beta-reductase activity which metabolized 3-dehydroecdysone (3DE) and 3-dehydro-20-hydroxyecdysone (3D20E) to ecdysone (E) and 20-hydroxyecdysone (20E), respectively, but had only very low amounts of ecdysone oxidase activity (E to 3DE) and no detectable ecdysone 20-monooxygenase activity (E to 20E). When the expression of the various genes was studied in the epidermis in vitro, 20E and 3D20E had similar effects, whereas E, 3DE, 26-hydroxyecdysone and 20,26-dihydroxyecdysone were ineffective. Exposure of Day 2 fifth-instar epidermis to 500 ng/ml of either 20E or 3D20E for 24 hr caused a rapid, biphasic increase in EcR-B1 mRNA. By contrast, EcR-A mRNA showed a less rapid initial increase followed by a slow steady rise and was less responsive to 3D20E. Ecdysone in a 1:1 mixture with 20E effectively halved the concentration of 20E needed to induce EcR-B1 mRNA but showed no synergism in the induction of EcR-A mRNA. The induction of MHR3 mRNA and of DDC mRNA in Day 2 fourth-instar epidermis as well as the suppression of DDC and LCP-14 gene expression by 3D20E was indistinguishable from that of 20E. Therefore, for Manduca larval epidermis, only 20E and 3D20E are biologically active ecdysteroids. Since the 3D20E can be converted to 20E by the epidermis, its effects are likely mediated by 20E.
为了确定哪些蜕皮甾类激素可能在烟草天蛾（Manduca sexta）幼虫表皮中具有生物活性，我们研究了几种已知的蜕皮甾类激素及其代谢产物对编码蜕皮激素受体（EcR）、烟草天蛾激素受体3（MHR3）、多巴脱羧酶（DDC）和一种幼虫表皮蛋白（LCP - 14）的基因表达的作用。二龄四龄幼虫和二龄五龄幼虫的表皮均含有显著的3β - 还原酶活性，该酶分别将3 - 脱氢蜕皮激素（3DE）和3 - 脱氢 - 20 - 羟基蜕皮激素（3D20E）代谢为蜕皮激素（E）和20 - 羟基蜕皮激素（20E），但蜕皮激素氧化酶活性（E转化为3DE）极低，且未检测到蜕皮激素20 - 单加氧酶活性（E转化为20E）。当在体外研究表皮中各种基因的表达时，20E和3D20E具有相似的作用效果，而E、3DE、26 - 羟基蜕皮激素和20,26 - 二羟基蜕皮激素则无作用效果。将二龄五龄幼虫表皮暴露于500 ng/ml的20E或3D20E中24小时，会导致EcR - B1 mRNA迅速出现双相增加。相比之下，EcR - A mRNA的初始增加速度较慢，随后缓慢稳定上升，且对3D20E的反应较小。与20E以1:1混合物形式存在的蜕皮激素有效地将诱导EcR - B1 mRNA所需的20E浓度减半，但在诱导EcR - A mRNA方面未显示出协同作用。3D20E对二龄四龄幼虫表皮中MHR3 mRNA和DDC mRNA以及对DDC和LCP - 14基因表达抑制的诱导作用与20E无法区分。因此对于烟草天蛾幼虫表皮，只有20E和3D20E是具有生物活性的蜕皮甾类激素。由于3D20E可被表皮转化为20E，其作用可能是由20E介导的。
