Fatty Acid Solubilizer from the Oral Disk of the Blowfly

BackgroundBlowflies are economic pests of the wool industry and potential vectors for epidemics. The establishment of a pesticide-free, environmentally friendly blowfly control strategy is necessary. Blowflies must feed on meat in order to initiate the cascade of events that are involved in reproduction including juvenile hormone synthesis, vitellogenesis, and mating. During feeding blowflies regurgitate salivary lipase, which may play a role in releasing fatty acids from triglycerides that are found in food. However, long-chain fatty acids show low solubility in aqueous solutions. In order to solubilize and ingest the released hydrophobic fatty acids, the blowflies must use a solubilizer.
MethodologyWe applied native PAGE, Edman degradation, cDNA cloning, and RT-PCR to characterize a protein that accumulated in the oral disk of the black blowfly, Phormia regina. In situ hybridization was carried out to localize the expression at the cellular level. A fluorescence competitive binding assay was used to identify potential ligands of this protein.
ConclusionA protein newly identified from P. regina (PregOBP56a) belonged to the classic odorant-binding protein (OBP) family. This gene was expressed in a cluster of cells that was localized between pseudotracheae on the oral disk, which are not accessory cells of the taste peg chemosensory sensilla that normally synthesize OBPs. At pH 7 and pH 6, PregOBP56a bound palmitic, stearic, oleic, and linoleic acids, that are mainly found in chicken meat. The binding affinity of PregOBP56a decreased at pH 5. We propose that PregOBP56a is a protein that solubilizes fatty acids during feeding and subsequently helps to deliver the fatty acids to the midgut where it may help in the process of reproduction. As such, PregOBP56a is a potential molecular target for controlling the blowfly.

研究背景：丽蝇是羊毛产业的经济害虫，同时也是传染病的潜在传播媒介。开发无农药、环境友好的丽蝇防控策略迫在眉睫。丽蝇必须取食肉类才能启动繁殖相关的一系列生理过程，包括保幼激素合成、卵黄发生以及交配行为。取食过程中，丽蝇会反刍唾液脂肪酶，该酶可能参与从食物中的甘油三酯中释放脂肪酸。然而，长链脂肪酸在水溶液中溶解度极低。为了溶解并摄取释放出的疏水性脂肪酸，丽蝇需要借助一种增溶剂。
研究方法：本研究采用非变性聚丙烯酰胺凝胶电泳（native PAGE）、埃德曼降解（Edman degradation）、cDNA克隆以及逆转录聚合酶链式反应（RT-PCR），对黑丽蝇（Phormia regina）口盘内富集的一种蛋白质进行表征。通过原位杂交技术在细胞水平上定位该蛋白的表达位点；采用荧光竞争性结合实验鉴定该蛋白的潜在配体。
研究结论：本研究从黑丽蝇（P. regina）中鉴定得到的新型蛋白PregOBP56a，属于经典的气味结合蛋白（odorant-binding protein, OBP）家族。该基因的表达定位于口盘伪气管之间的细胞簇，这类细胞并非通常合成气味结合蛋白的味觉毛化学感应感器的附属细胞。在pH 7和pH 6条件下，PregOBP56a可结合主要存在于鸡肉中的棕榈酸、硬脂酸、油酸以及亚油酸；在pH 5条件下，PregOBP56a的结合亲和力显著下降。本研究推测，PregOBP56a是一种在取食过程中溶解脂肪酸，并协助将脂肪酸转运至中肠的蛋白，该过程可能参与丽蝇的繁殖活动。因此，PregOBP56a可作为防控丽蝇的潜在分子靶标。