小柯机器人

美国波士顿儿童医院Min Dong等研究人员合作发现，果蝇细胞中的CRISPR筛选将Vsg鉴定为Tc毒素受体。这一研究成果于2022年9月28日在线发表在国际学术期刊《自然》上。

研究人员在黑腹果蝇S2R+细胞中使用全基因组CRISPR-Cas9介导的敲除筛选，并确定Vsgun（Vsg）是典型的Photorhabdus luminescens Tc毒素（pTc）的一个受体。该毒素能识别Vsg的细胞外O-糖基化粘蛋白样结构域，该结构域含有脯氨酸、苏氨酸和丝氨酸的高密度重复序列（HD-PTS）。蚊子和甲虫的Vsg同源物含有HD-PTS，可作为pTc受体，而没有HD-PTS的同源物，如飞蛾和人类，不是pTc受体。Vsg在免疫细胞中表达，包括血细胞和脂肪体细胞。Vsg基因敲除果蝇的血细胞对pTc有抵抗力，并在有pTc的情况下保持吞噬作用，通过蚊子Vsg的转基因表达，它们对pTc的敏感性得到恢复。最后，Vsg基因敲除的果蝇显示出细菌负荷和P. luminescens感染致死率的减少。这些研究结果确定了一个蛋白型Tc毒素受体，揭示了Tc毒素如何促进P. luminescens的致病机制，并建立了一个全基因组CRISPR筛选方法来研究杀虫毒素和病原体。

据介绍，昆虫病原体线虫被广泛用作生物杀虫剂。它们的杀虫活性取决于共生细菌，如P. luminescens，它产生毒素复合体（Tc）毒素作为主要毒力因子。没有任何Tc毒素的蛋白受体是已知的，这限制了人们对其特异性和致病机制的理解。

附：英文原文

Title: CRISPR screens in Drosophila cells identify Vsg as a Tc toxin receptor

Author: Xu, Ying, Viswanatha, Raghuvir, Sitsel, Oleg, Roderer, Daniel, Zhao, Haifang, Ashwood, Christopher, Voelcker, Cecilia, Tian, Songhai, Raunser, Stefan, Perrimon, Norbert, Dong, Min

Issue&Volume: 2022-09-28

Abstract: Entomopathogenic nematodes are widely used as biopesticides1,2. Their insecticidal activity depends on symbiotic bacteria such as Photorhabdus luminescens, which produces toxin complex (Tc) toxins as major virulence factors3,4,5,6. No protein receptors are known for any Tc toxins, which limits our understanding of their specificity and pathogenesis. Here we use genome-wide CRISPR–Cas9-mediated knockout screening in Drosophila melanogaster S2R+ cells and identify Visgun (Vsg) as a receptor for an archetypal P.luminescens Tc toxin (pTc). The toxin recognizes the extracellular O-glycosylated mucin-like domain of Vsg that contains high-density repeats of proline, threonine and serine (HD-PTS). Vsg orthologues in mosquitoes and beetles contain HD-PTS and can function as pTc receptors, whereas orthologues without HD-PTS, such as moth and human versions, are not pTc receptors. Vsg is expressed in immune cells, including haemocytes and fat body cells. Haemocytes from Vsg knockout Drosophila are resistant to pTc and maintain phagocytosis in the presence of pTc, and their sensitivity to pTc is restored through the transgenic expression of mosquito Vsg. Last, Vsg knockout Drosophila show reduced bacterial loads and lethality from P.luminescens infection. Our findings identify a proteinaceous Tc toxin receptor, reveal how Tc toxins contribute to P.luminescens pathogenesis, and establish a genome-wide CRISPR screening approach for investigating insecticidal toxins and pathogens.

DOI: 10.1038/s41586-022-05250-7

Source: https://www.nature.com/articles/s41586-022-05250-7

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