Evidence that microRNAs are part of the molecular toolkit regulating adult reproductive diapause in the mosquito, Culex pipiens

For many insects, diapause is the primary mechanism for surviving unfavorable seasons. Some aspects of diapause regulation are well known, but we still lack a mechanistic understanding of molecular mechanisms that control the diapause pathway. Accumulating evidence suggests microRNAs regulate diapause in evolutionarily diverse insect species including flesh flies and moths, and, it is likely that microRNAs regulate multiple characteristics of diapause, including arrested egg follicle development and fat hypertrophy, in females of the Northern house mosquito, Culex pipiens. To investigate microRNA regulation of diapause in this species, we measured changes in egg follicle development and total lipid content over 22 days following adult emergence. We also evaluated changes in the abundance of candidate microRNAs associated with these physical changes during the same time frame. We found egg follicle size and lipid content were nearly the same in diapausing and nondiapausing females on the day of adult emergence, and then diverged over time such that by day 22 diapausing females had significantly smaller egg follicles and higher total lipids than their nondiapausing counterparts. Several microRNAs associated with lipid metabolism in insects, including miR-14-3p, miR-277-3p, and miR-305-5p, were underexpressed in diapausing females compared to nondiapausing females on the day of adult emergence, which suggests microRNA regulation occurs ahead of observed changes in these two features of the diapause phenotype. We also found miR-309-3p, miR-375-3p which stimulate ovarian development in other mosquito species, were underexpressed in diapausing females of Cx. pipiens at times after diapause is fully established and may be responsible for the arrest in ovarian development in this species. Taken together, our results demonstrate that changes in the abundance of some microRNAs is associated with phenotypic changes in diapause Cx. pipiens and suggests this epigenetic mechanism is part of the molecular toolkit regulating diapause.

对于多数昆虫而言，滞育（diapause）是抵御不良季节的核心生存机制。尽管滞育调控的部分环节已得到充分阐明，但我们对调控滞育通路的分子机制仍缺乏机制性认知。越来越多的证据表明，微小RNA（microRNAs，miRNAs）可调控进化上多样的昆虫物种的滞育过程，其中包括肉蝇与蛾类；而在尖音库蚊（Culex pipiens）的雌性个体中，微小RNA或可调控滞育的多项特征，包括停滞的卵泡发育与脂肪肥大。为探究该物种滞育过程中的微小RNA调控机制，我们检测了成虫羽化后22天内卵泡发育与总脂含量的动态变化，并同步评估了与这些生理变化相关的候选微小RNA的丰度改变。结果显示，成虫羽化当日，滞育与非滞育雌性个体的卵泡大小与脂含量基本一致；随后二者的相关指标出现分化，至第22天时，滞育雌性的卵泡显著小于非滞育个体，而总脂含量则显著更高。相较于非滞育雌性，成虫羽化当日的滞育雌性体内，多种与昆虫脂质代谢相关的微小RNA——包括miR-14-3p、miR-277-3p与miR-305-5p——表达量显著下调，这提示微小RNA的调控先于这两个滞育表型特征的变化发生。我们还发现，在其他蚊种中可促进卵巢发育的miR-309-3p与miR-375-3p，在滞育完全建立后的尖音库蚊滞育雌性体内表达量下调，这或为该物种卵巢发育停滞的成因。综上，本研究结果表明，部分微小RNA的丰度变化与滞育尖音库蚊的表型改变显著相关，提示这种表观遗传机制是调控滞育的分子工具库的组成部分。