Hemocyanin is a multifunctional respiratory glycoprotein, which has also been implicated in other biological functions in shrimp. Moreover, recent studies have revealed that hemocyanin is also involved in a broad range of immune-related activities in shrimp. However, in spite of the considerable interest in unraveling the reasons behind the multiple immune-related functions of hemocyanin, little is known about its transcriptional regulation. Here, DNA pull-down and Liquid Chromatography - Tandem Mass Spectrometry (LC-MS/MS) analyses were used to isolate and identify the putative transcription factor(s) that are involved in the transcriptional regulation of the small subunit hemocyanin gene of Penaeus vannamei (PvHMCs). Krüppel-like factor (designated PvKruppel), a zinc finger transcription factor homolog in P. vannamei, was identified among the putative transcription factors, while bioinformatics analysis revealed the presence of Krüppel-like factor binding site (KLF motif) on the core promoter region of PvHMCs. Mutational analysis and electrophoretic mobility shift assay (EMSA) confirmed that PvKruppel could bind to the KLF motif on the core promoter region of PvHMCs. Moreover, in response to lipopolysaccharide (LPS), Vibrio parahaemolyticus and white spot syndrome virus (WSSV) challenge, transcript levels of PvKruppel and PvHMCs were negatively correlated. Furthermore, overexpression of PvKruppel significantly reduced the promoter activity of PvHMCs, while PvKruppel knockdown by RNA interference or lipopolysaccharides (LPS) stimulation resulted in a significant increase in the transcript level of PvHMCs. Taken together, our present study provides mechanistic insights into the transcriptional regulation of PvHMCs by PvKruppel during shrimp immune response to pathogens.

血蓝蛋白是一种多功能的呼吸糖蛋白，也与虾的其他生物学功能有关。此外，最近的研究表明，血蓝蛋白还参与了虾的多种免疫相关活动。然而，尽管人们对揭示血蓝蛋白的多种免疫相关功能背后的原因十分感兴趣，但对其转录调控的了解却很少。在这里，使用DNA下拉和液相色谱-串联质谱（LC-MS / MS）分析来分离和鉴定与南美白对虾小亚基血蓝蛋白基因的转录调控有关的推定转录因子。（PvHMC）。Krüppel样因子（称为PvKruppel），是锌指转录因子的同源物在假定的转录因子中鉴定出南美白对虾，而生物信息学分析显示，PvHMCs核心启动子区域存在Krüppel样因子结合位点（KLF基序）。突变分析和电泳迁移率变动分析（EMSA）证实PvKruppel可以与PvHMCs核心启动子区域的KLF基序结合。此外，响应脂多糖（LPS），副溶血性弧菌和白斑综合症病毒（WSSV）攻击，PvKruppel和PvHMCs的转录水平呈负相关。此外，PvKruppel的过表达显着降低了PvHMCs的启动子活性，而RNA干扰或脂多糖（LPS）刺激引起的PvKruppel敲低导致PvHMCs的转录水平显着增加。综上所述，我们的当前研究提供了对虾对病原体的免疫应答过程中PvKruppel对PvHMCs转录调控的机制性见解。