姜黄素调控转录因子FOXP3 影响HIV-1 感染辅助受体CCR5 的作用机制 研究.

Objective: To investigate the mechanism of curcumin affecting HIV-1 infection co-receptor CCR5 by regulating transcription factor FOXP3. Methods: Binding sites of transcription factor FOXP3 on CCR5 promoter were predicted and analyzed by bioinformatics method. AutoDock 4.2 software was used to connect curcumin and FOXP3 flexibly. MTT assay was used to detect cytotoxcity of curcumin on activity of Jurkat cells. qRT-PCR and Western blot were used to detect expression levels of CCR5 and FOXP3 mRNA and protein in Jurkat cells that were treated with different concentrations of curcumin. pcDNA3.1-FOXP3 expression vector was built and combined with the prediction results of transcription factors. The mutant CCR5 gene fragment was amplified by Overlap PCR, and the mutant CCR5 promoter recombinant vector pFireRluc-Mt-CCR5 was constructed. Binding site between transcription factor FOXP3 and CCR5 promoter was verified by double luciferase reporter gene assay. Results: Results of JASPAR transcription factor prediction showed that there was a binding site between CCR5 promoter and transcription factor FOXP3; molecular docking results showed that curcumin could bind to the active region of FOXP3; MTT results showed that curcumin inhibited the activity of Jurkat cells after 24 hours, and the IC50 was 34.48 μmol/L. qRT-PCR and Western blot showed that expression levels of CCR5 and FOXP3 mRNA and protein were decreased in a dose-dependent manner after different concentrations of curcumin treated Jurkat cells; double luciferase reporter gene confirmed that FOXP3 could bind to CCR5 promoter, and the transcription factor FOXP3 could regulate the activity of CCR5 promoter; results of the recovery experiment of FOXP3 on curcumin showed that when the curcumin concentration was 60 μmol/L, relative value of luciferase activity in HEK293T cells with pcDNA3.1-FOXP3 and pFireRluc-Wt-CCR5 was significantly higher than that in pFireRluc-Wt-CCR5+curcumin-60 group （P<0.01）. Conclusion: FOXP3 can regulate the activity of CCR5 promoter, and the mechanism may be that curcumin affects activity of CCR5 promoter by acting on binding site of FOXP3 and CCR5 promoter. [ABSTRACT FROM AUTHOR]