The mosquito transcriptome is shaped by the individual and interactive effects of warmer temperature and aging

In mosquitoes, warmer temperature accelerates the progression of senescence, weakening their ability to survive and respond to an infection. Thus, we hypothesized that this phenomenon is driven by changes in gene expression. To test this hypothesis, we employed an RNA-sequencing approach to investigate how warmer temperature and aging, individually and interactively, shape the transcriptome of the female African malaria mosquito, Anopheles gambiae. Warmer temperature increases the expression of genes involved in metabolism and cell signaling but decreases the expression of genes involved in replication, transcription, translation, and protein processing. Aging decreases the expression of metabolism-related genes but increases the expression of protein processing and transcriptional genes. However, early and late aging have opposing effects on many biological processes, with 1-day-old mosquitoes having a largely different transcriptomic profile than older mosquitoes. Importantly, warmer temperature and aging interactively shape gene expression patterns, where, for example, aging-dependent changes in immune gene expression accelerate at warmer temperatures. Altogether, warmer temperature and aging, individually and interactively, alter the mosquito's transcriptomic profile, reflecting changes in their ability to survive and overcome an infection. Overall design: We set out to investigate how warmer temperature and aging, individually and interactively, shape the transcriptome of naïve and immune-induced mosquitoes. To do so, we reared Anopheles gambiae (Giles sensu stricto) at 27°C, 30°C or 32°C, and allocated adult females that were 1, 5, 10, and 15 days post-eclosion to two immune treatment groups: (i) naïve and (ii) immune induced by intrathoracic injection of heat-killed Escherichia coli. At 6 h post immune treatment we isolated RNA from pools of 15 whole-body female mosquitoes for each temperature-age combination. We completed 5 independent biological trials, resulting in 120 samples derived from 1772 mosquitoes. We then conducted RNA-sequencing and analysis to identify how immune induction, warmer temperature, aging, and the interaction between warmer temperature and aging, alter gene expression profiles.

在蚊虫中，温度升高会加速衰老进程，削弱其生存能力与感染应答能力。据此，我们提出假说：该现象由基因表达变化所驱动。为验证这一假说，我们采用RNA测序（RNA-sequencing）技术，探究升温与衰老单独及共同作用如何塑造雌性非洲疟疾蚊虫——冈比亚按蚊（Anopheles gambiae）的转录组。升温会上调代谢与细胞信号通路相关基因的表达，却下调复制、转录、翻译及蛋白质加工相关基因的表达。衰老则会下调代谢相关基因的表达，同时上调蛋白质加工与转录相关基因的表达。不过，衰老早期与晚期对诸多生物学过程的影响截然相反：羽化后1日龄蚊虫的转录组特征与老龄蚊虫存在显著差异。值得注意的是，升温与衰老可共同调控基因表达模式，例如，免疫相关基因的衰老依赖性表达变化在升温条件下会加速。总而言之，升温与衰老分别及联合作用均可改变蚊虫的转录组特征，反映出其生存与抗感染能力的变化。整体实验设计：本研究旨在探究升温与衰老单独及共同作用如何塑造未免疫与免疫诱导蚊虫的转录组。为此，我们将冈比亚按蚊（严格指名亚种，Giles sensu stricto）分别饲养于27℃、30℃或32℃环境中，并将羽化后1、5、10、15日龄的成年雌蚊分为两组免疫处理组：（i）未免疫组；（ii）经胸内注射热灭活大肠杆菌（Escherichia coli）诱导免疫组。免疫处理6小时后，我们从每个温度-年龄组合的15只整身雌蚊混合样本中提取RNA。本研究共完成5次独立生物学重复，最终从1772只蚊虫中获得120个样本。随后我们开展RNA测序及分析，以明确免疫诱导、升温、衰老以及升温与衰老的交互作用如何改变基因表达谱。