National Engineering Laboratory for Modern Silk, Department of Applied Biology, School of Biology and Basic Medical Sciences, Medical College, Soochow University, Suzhou, China.
Arch Insect Biochem Physiol. 2012 Mar;79(3):182-94. doi: 10.1002/arch.21019.
Diapause in Bombyx mori eggs is induced by temperature and photoperiod at the stage of embryonic development in the maternal generation. In those diapause eggs, Esterase-A4 is suggested to serve as a diapause-termination timer (TIME-EA4), because its ATPase activity shows an interval-timer elevation after acid treatment or chilling of eggs to break diapause. To clarify whether the timed ATPase activity of TIME-EA4 is related to its gene (Ea4) expression, we analyzed Ea4 mRNA of eggs in diapause-inducing environmental conditions. Reverse transcription PCR (RT-PCR) analysis showed that the level of Ea4 mRNA was lower in 15DD than in 25LL (P < 0.01) or in 20LD (P < 0.01) but did not oscillate when photoperiod and temperature periodically oscillated (P > 0.05). Furthermore, expressed sequence tag profile and gene microarray analysis demonstrated that Ea4 showed stage-specific and tissue-specific expression during postembryonic stages, high Ea4 mRNA in the spinning and eclosion stages, and in integument and head, but low in gonads of fifth-instar day-3 larvae. Then we analyzed the relationship between TIME-EA4 ATPase activities and Ea4 gene expression. The ATPase activities in diapause eggs laid by the resultant adults transiently elevated after treatments to break diapause, that is, at 1.5 h after common-acid treatment and day 13 after chilling at 5°C. However, these elevations of enzyme activities were not accompanied by any increases in Ea4 mRNA levels. In conclusion, the termination of the Bombyx embryonic diapause is related to TIME-EA4 but not to its gene expression.
家蚕卵滞育是由母代胚胎发育期的温度和光周期诱导的。在这些滞育卵中，酯酶-A4 被认为是滞育终止定时器（TIME-EA4），因为其 ATP 酶活性在酸处理或冷藏卵以打破滞育后显示出间隔定时器升高。为了阐明 TIME-EA4 的定时 ATP 酶活性是否与其基因（Ea4）表达有关，我们分析了在诱导滞育的环境条件下卵中的 Ea4 mRNA。逆转录 PCR（RT-PCR）分析表明，在 15DD 中 Ea4 mRNA 的水平低于 25LL（P<0.01）或 20LD（P<0.01），但在光周期和温度周期性波动时没有波动（P>0.05）。此外，表达序列标签谱和基因微阵列分析表明，Ea4 在胚胎后阶段表现出阶段特异性和组织特异性表达，在吐丝和羽化阶段以及在表皮和头部中表达水平较高，但在第五龄第三天的性腺中表达水平较低。然后，我们分析了 TIME-EA4 ATP 酶活性与 Ea4 基因表达之间的关系。在打破滞育的处理后，即酸处理后 1.5 小时和 5°C 冷藏后第 13 天，由成虫产下的滞育卵中的 ATP 酶活性会短暂升高。然而，这些酶活性的升高并没有伴随着 Ea4 mRNA 水平的任何增加。总之，家蚕胚胎滞育的终止与 TIME-EA4 有关，而与基因表达无关。
