Responses of Gene to Infection and Thermal Stress and Its Transcriptional Regulation Analysis in .

Heat-shock protein 70 (HSP70) is a molecular chaperone that plays critical roles in cell protein folding and metabolism, which helps to protect cells from unfavorable environmental stress. is one of the most important economic breeding species in the coastal provinces of south China. To date, the expression and transcriptional regulation of in () has not been well characterized. In this study, the expression levels of gene in different tissues and different stress conditions were detected. The results showed that the gene was ubiquitously expressed in sampled tissues and was the highest in hepatopancreas, followed by hemocytes. In hepatopancreas and hemocytes, the gene was significantly up-regulated by infection, thermal stress, and combined stress ( infection and thermal stress combination), indicating that HSP70 is involved in the stress response and the regulation of innate immunity. Furthermore, a 2383 bp of 5'-flanking region sequence of the gene was cloned, and it contains a presumed core promoter region, a CpG island, a (TG) simple sequence repeat (SSR), and many potential transcription factor binding sites. The activity of promoter was evaluated by driving the expression of luciferase gene in HEK293FT cells. A series of experimental results indicated that the core promoter region is located between -189 bp and +46 bp, and high-temperature stress can increase the activity of promoter. Sequence-consecutive deletions of the luciferase reporter gene in HEK293FT cells revealed two possible promoter activity regions. To further identify the binding site of the key transcription factor in the two regions, two expression vectors with site-directed mutation were constructed. The results showed that the transcriptional activity of NF-1 site-directed mutation was significantly increased ( < 0.05), whereas the transcriptional activity of NF-κB site-directed mutation was significantly reduced. These results suggest that NF-1 and NF-κB may be two important transcription factors that regulate the expression of gene.