Data from: Engineered action at a distance: blood-meal-inducible paralysis in Aedes aegypti

Background: Population suppression through mass-release of Aedes aegypti
males carrying dominant-lethal transgenes has been demonstrated in the
field. Where population dynamics show negative density-dependence,
suppression can be enhanced if lethality occurs after the
density-dependent (i.e. larval) stage. Existing molecular tools have
limited current examples of such Genetic Pest Management (GPM) systems to
achieving this through engineering ‘cell-autonomous effectors’ i.e. where
the expressed deleterious protein is restricted to the cells in which it
is expressed – usually under the control of the regulatory elements (e.g.
promoter regions) used to build the system. This limits the flexibility of
these technologies as regulatory regions with useful spatial, temporal or
sex-specific expression patterns may only be employed if the cells they
direct expression in are simultaneously sensitive to existing effectors,
and also precludes the targeting of extracellular regions such as
cell-surface receptors. Expanding the toolset to ‘non-cell autonomous’
effectors would significantly reduce these limitations.
Methodology/Principal Findings: We sought to engineer female-specific,
late-acting lethality through employing the Ae. aegypti VitellogeninA1
promoter to drive blood-meal-inducible, fat-body specific expression of
tTAV. Initial attempts using pro-apoptotic effectors gave no evident
phenotype, potentially due to the lower sensitivity of
terminally-differentiated fat-body cells to programmed-death signals.
Subsequently, we dissociated the temporal and spatial expression of this
system by engineering a novel synthetic effector (Scorpion neurotoxin –
TetO-gp67.AaHIT) designed to be secreted out of the tissue in which it was
expressed (fat-body) and then affect cells elsewhere (neuro-muscular
junctions). This resulted in a striking, temporary-paralysis phenotype
after blood-feeding. Conclusions/Significance: These results are
significant in demonstrating for the first time an engineered ‘action at a
distance’ phenotype in a non-model pest insect. The potential to
dissociate temporal and spatial expression patterns of useful endogenous
regulatory elements will extend to a variety of other pest insects and
effectors.

背景：通过大规模释放携带显性致死转基因（dominant-lethal transgenes）的埃及伊蚊（Aedes aegypti）雄蚊来抑制种群数量的方法已在实地得到验证。在种群动态呈现负密度依赖（negative density-dependence）的情况下，若致死效应发生在密度依赖阶段（即幼虫阶段（larval stage））之后，抑制效果可得到增强。现有分子工具限制了此类遗传害虫管理（Genetic Pest Management, GPM）系统的现有实例，使其仅能通过改造‘细胞自主效应因子（cell-autonomous effectors）’来实现这一目标——即表达的有害蛋白被限制在其表达的细胞内，通常受构建系统所用调控元件（regulatory elements）（如启动子区域（promoter regions））的控制。这限制了这些技术的灵活性，因为只有当调控区域所引导表达的细胞同时对现有效应因子敏感时，具有有用空间（spatial）、时间（temporal）或性别特异性（sex-specific）表达模式的调控区域才能被利用；此外，这也排除了靶向细胞外区域（extracellular regions）（如细胞表面受体（cell-surface receptors））的可能性。将工具集扩展至‘非细胞自主效应因子（non-cell autonomous effectors）’将显著减少这些限制。

方法/主要发现：我们尝试通过利用埃及伊蚊卵黄蛋白原A1启动子（VitellogeninA1 promoter）驱动tTAV的吸血诱导性（blood-meal-inducible）、脂肪体特异性（fat-body specific）表达，来改造雌性特异性、晚期作用的致死效应。最初使用促凋亡效应因子（pro-apoptotic effectors）的尝试未产生明显表型，这可能是由于终末分化的脂肪体细胞对程序性死亡信号（programmed-death signals）的敏感性较低。随后，我们通过改造一种新型合成效应因子（synthetic effector）（蝎神经毒素（Scorpion neurotoxin）——TetO-gp67.AaHIT）来分离该系统的时间和空间表达；这种效应因子被设计为从其表达的组织（脂肪体）中分泌出来，然后影响其他部位的细胞（神经肌肉接头（neuro-muscular junctions））。这导致吸血后出现显著的暂时性麻痹表型（temporary-paralysis phenotype）。

结论/意义：这些结果具有重要意义，首次在非模式害虫（non-model pest insect）中证明了改造的‘远距离作用表型（action at a distance phenotype）’。分离有用内源性调控元件（endogenous regulatory elements）的时间和空间表达模式的潜力将扩展到多种其他害虫和效应因子。