Aedes aegypti is the principle vector of many arboviruses, including dengue virus and Zika virus, which are transmitted when an infected female mosquito takes a blood meal in order to initiate vitellogenesis. During blood digestion, ~ 10 mM heme-iron is ingested into the midgut lumen. While heme acts as both a nutrient and signaling molecule during blood digestion, it can also be highly toxic if left unchaperoned. Both signaling by, and degradation of, heme are intracellular processes, occurring in the nucleus and cytoplasm, respectively. However, the precise mechanism of heme uptake into the midgut epithelium is not currently known. We used next generation RNA sequencing with the goal to identify genes that code for membrane bound heme import protein(s) responsible for heme uptake into the midgut epithelium. Heme deprivation increased uptake of a heme fluorescent analog in cultured cells, while treatment of midguts with an excess of heme decreased uptake, confirming physiological changes were occurring in these heme-sensitive cells/tissues prior to sequencing. A list of candidate genes was assembled for each of the experimental sample sets, which included Aag2 and A20 cultured cells as well as midgut tissue, based on the results of a differential expression analysis, soft cluster analysis and number of predicted transmembrane domains. Lastly, the functions related to heme transport were examined through RNAi knockdown. Despite a large number of transmembrane domain containing genes differentially expressed in response to heme, very few were highly differentially expressed in any of the datasets examined. RNAi knockdown of a subset of candidates resulted in subtle changes in heme uptake, but minimal overall disruption to blood digestion/egg production. These results could indicate that heme import in Ae. aegypti may be controlled by a redundant system of multiple distinct transport proteins. Alternatively, heme membrane bound transport in Ae. aegypti could be regulated post-translationally.

中文翻译：

---

埃及白纹伊蚊培养的细胞和离体中肠对血红素过量或不足的转录组学分析：寻求转录调节的血红素转运蛋白。

埃及伊蚊是许多虫媒病毒（包括登革热病毒和寨卡病毒）的主要载体，当感染的雌性蚊子吸食血粉以启动卵黄体形成时，便会传播这种病毒。在血液消化过程中，约10 mM的血红素铁被摄入中肠腔。虽然血红素在血液消化过程中既是营养物质又是信号分子，但如果不进行陪护，它也可能具有剧毒。血红素的信号传导和降解都是细胞内过程，分别发生在细胞核和细胞质中。然而，目前尚不知道血红素摄取到中肠上皮的确切机制。我们使用了下一代RNA测序技术，目的是鉴定编码膜结合的血红素导入蛋白的基因，这些蛋白负责将血红素摄取到中肠上皮中。血红素剥夺增加了培养细胞中血红素荧光类似物的摄取，而用过量血红素处理中肠会降低摄取，这证实了测序之前这些血红素敏感细胞/组织中发生了生理变化。根据差异表达分析，软聚类分析和预测的跨膜结构域的结果，为每个实验样品集（包括Aag2和A20培养的细胞以及中肠组织）装配了候选基因列表。最后，通过RNAi敲除检查了与血红素转运有关的功能。尽管有大量的跨膜结构域包含响应血红素而差异表达的基因，但在所检查的任何数据集中，很少有高度差异表达的基因。候选子集的RNAi敲除导致血红素摄取的细微变化，但对血液消化/鸡蛋产生的总体破坏最小。这些结果可能表明血红素进口到Ae。埃及可能由多个不同转运蛋白的冗余系统控制。备选地，血红素膜结合在Ae中的运输。可在翻译后对埃及埃及进行监管。