肽聚糖识别蛋白PGRP-SC在家蝇肠道免疫中的功能分析

Functional analysis of peptidoglycan recognition protein PGRP-SC in intestinal immunity of Musca domestica

【目的】探究家蝇Musca domestica幼虫肽聚糖识别蛋白PGRP-SC在应对肠道入侵细菌中的作用。【方法】通过投喂表达Md PGRP-SC dsRNA的宿主大肠杆菌Escherichia coli干扰家蝇初孵幼虫Md PGRP-SC基因表达,并利用q PCR技术检测干扰效果;在荧光显微镜下观察干扰后试虫肠道内表达绿色荧光蛋白(green fluorescent protein,GFP)的大肠杆菌存活情况,同时通过比色法检测干扰后试虫肠道组织内的H2O2含量,利用q PCR检测试虫肠道内抗菌肽基因的表达情况。【结果】q PCR结果显示,投喂干扰24 h后家蝇幼虫体内Md PGRP-SC基因的相对表达量显著降低。对PGRP-SC干扰组和GFP对照组试虫投喂表达GFP的大肠杆菌,30 min后干扰组家蝇肠道内荧光亮度弱于GFP对照组。同时,干扰组试虫肠道内H2O2含量与空白对照或GFP对照组相比均没有显著差异,而干扰组中抗菌肽表达水平显著高于GFP对照组。【结论】Md PGRP-SC在控制家蝇幼虫肠道免疫敏感程度中起作用。

【Aim 】 The objective of this research is to investigate the immunological functions of peptidoglycan recognition protein SC in the housefly,Musca domestica,in response to invading bacteria.【Methods】We knocked down the expression level of Md PGRP-SC by feeding the newly-hatched larvae of M. domestica with Escherichia coli expressing double-strand RNA and then measured the interference effect with quantitative real-time PCR（ q PCR）. The treated larvae were fed with GFP-expressing E.coli,and then the tempo-spatial dynamics of these ingested bacteria was observed under fluorescent microscope. The hydrogen peroxide content and the expression profiles of antimicrobial peptide genes in the intestine of the larvae fed with dsRNA were tested by colorimetry and q PCR methods,respectively.【Results】The q PCR results showed a significant decrease in the relative expression level of PGRP-SC in the interference group. After M. domestica larvae fed with E. coli expressing GFP,a rapid fadeout of fluorescence in the Md PGRP-SC interference group was observed compared with the GFP control group.The hydrogen peroxide content in each group had no significant change,while the expression levels of all the four AMP genes were up-regulated dramatically in the gut of Md PGRP-SC depleted larvae.【Conclusion】Md PGRP-SC plays a role in regulating the sensitivity of intestinal immune reaction in housefly larvae.