Zhang Shuang, Qiu Wei, Chen Yong-gui, Yuan Feng-Hua, Li Chao-Zheng, Yan Hui, Weng Shao-Ping, He Jian-Guo
MOE Key Laboratory of Aquatic Product Safety/State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, 135 Xingang Road West, Guangzhou 510275, PR China.
School of Marine Sciences, Sun Yat-sen University, 135 Xingang Road West, Guangzhou 51027, PR China.
Fish Shellfish Immunol. 2015 Feb;42(2):413-25. doi: 10.1016/j.fsi.2014.10.023. Epub 2014 Oct 27.
Flightless-I (FliI) is a protein negatively modulates the Toll-like receptor (TLR) pathway through interacting with Myeloid differentiation factor 88 (MyD88). To investigate the function of FliI in innate immune responses in invertebrates, Litopenaeus vannamei FliI (LvFliI) was identified and characterized. The full-length cDNA of LvFliI is 4, 304 bp long, with an open reading frame (ORF) encoding a putative protein of 1292 amino acids, including 12 leucine-rich repeat (LRR) domains at the N-terminus and 6 gelsolin homology (GEL) domains at the C-terminus. The LvFliI protein was located in the cytoplasm and LvFliI mRNA was constitutively expressed in healthy L. vannamei, with the highest expression level in the muscle. LvFliI could be up-regulated in hemocytes after lipopolysaccharide (LPS), poly I:C, CpG-ODN2006, Vibrio parahaemolyticus, Staphylococcus aureus, and white spot syndrome virus (WSSV) challenges, suggesting a stimulation response of LvFliI to bacterial and immune stimulant challenges. Upon LPS stimulation, overexpression of LvFliI in Drosophila Schneider 2 cells led to downregulation of Drosophila and shrimp antimicrobial peptide (AMP) genes. Knockdown of LvFliI by RNA interference (RNAi) resulted in an increase of the expression of three shrimp AMP genes (PEN2, crustin, and Lyz1). However, the mortality rates of LvFliI-knockdown shrimp in response to V. parahaemolyticus, S. aureus or WSSV infections were not significantly different from those of the control group. Taken together, all the results suggested that LvFliI may play a negative role in TLR signaling response in L. vannamei.
非飞行蛋白I（FliI）是一种通过与髓样分化因子88（MyD88）相互作用来负向调节Toll样受体（TLR）信号通路的蛋白质。为了研究FliI在无脊椎动物先天免疫反应中的功能，我们对凡纳滨对虾FliI（LvFliI）进行了鉴定和表征。LvFliI的全长cDNA长4304 bp，有一个开放阅读框（ORF），编码一个推定的含1292个氨基酸的蛋白质，其N端包括12个富含亮氨酸重复序列（LRR）结构域，C端有6个凝溶胶蛋白同源（GEL）结构域。LvFliI蛋白定位于细胞质中，LvFliI mRNA在健康的凡纳滨对虾中组成性表达，在肌肉中的表达水平最高。在脂多糖（LPS）、聚肌苷酸:聚胞苷酸（poly I:C）、CpG-ODN2006、副溶血性弧菌、金黄色葡萄球菌和白斑综合征病毒（WSSV）刺激后，血细胞中的LvFliI可被上调，这表明LvFliI对细菌和免疫刺激物刺激有应激反应。在LPS刺激下，LvFliI在果蝇Schneider 2细胞中的过表达导致果蝇和对虾抗菌肽（AMP）基因的下调。通过RNA干扰（RNAi）敲低LvFliI导致三种对虾AMP基因（PEN2、抗真菌肽和Lyz1）的表达增加。然而，敲低LvFliI的对虾在应对副溶血性弧菌、金黄色葡萄球菌或WSSV感染时的死亡率与对照组没有显著差异。综上所述，所有结果表明LvFliI可能在凡纳滨对虾的TLR信号反应中起负向作用。
