Journal: PLOS Pathogens

Article Title: YAP Inhibits HIV-1 transcription and promotes HIV-1 latency by regulating E3 ubiquitin ligase UHRF1 mediated tat degradation

doi: 10.1371/journal.ppat.1013906

Figure Lengend Snippet: A) Differential expression of the YAP gene in both AIDS patients and healthy controls (HCs) in the GEO database ( GSE140713 ) was analyzed via the GEO2R online tool. B) Immunoblot analysis (top) showing the levels of YAP expression in TZM-bl transfected with the increased amounts of p cDNA3.1-YAP-Flag plasmid. HIV-1 LTR-driven luciferase activity in HIV-1 IIIB -infected TZM-bl cells transfected with the indicated amounts of p cDNA3.1-YAP-Flag is shown in the bottom. C) Immunoblot analysis (top) showing the levels of Flag-tag expression in Jurkat T cells stably transduced with lentivirus. The band indicated by the arrow is the Flag-tag species. Jurkat T cells stably expressing YAP-Flag or the lentiviral vector were infected with HIV-1 IIIB, and the p24 production of the supernatants was detected by ELISA (bottom). D) RT-qPCR analysis of the YAP knockdown efficiency in Jurkat T cells generated by a doxycycline (dox)-inducible Tet-On system (CTGF, a classic YAP target gene) following treatment of Jurkat T cells with the HIV-1 IIIB virus with or without dox for 48 h. E , F) The p24 production and a luciferase activity were measured by ELISA and a luciferase assay. G) Immunoblot analysis (top) showing the levels of YAP expression in hPBMCs isolated from HIV-negative participants transduced with shRNA targeting YAP. The p24 production in the supernatant of HIV-1 IIIB infected hPBMCs was shown in the bottom. H) Immunoblot analysis (top) showing the levels of YAP expression in TZM-bl transfected with the indicated siRNAs. HIV-1 LTR-driven luciferase activity in HIV-1 IIIB -infected TZM-bl cells transfected with siRNAs targeting YAP and TAZ was shown in the bottom. I) RT-qPCR analysis of the expression of HIV-1 transcripts at different sites (initiation, proximal (Pro), intermediate (Int), and distal (Dis)) of HIV-1 IIIB infected Jurkat T cells with or without Dox for 48 h. Data were normalized to the values of the control group and were presented as the means ± SDs. To detect significant differences, one-way ANOVA was conducted in panels A) , B) , and H) , and two-way ANOVA was conducted in panels C) , D) , E) , F) , G) , and I) (* P < 0.05, ** P < 0.01, *** P < 0.001).

Article Snippet: CPI-203 and doxycycline were purchased from MedChemExpress.

Techniques: Quantitative Proteomics, Western Blot, Expressing, Transfection, Plasmid Preparation, Luciferase, Activity Assay, Infection, FLAG-tag, Stable Transfection, Transduction, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Knockdown, Generated, Virus, Isolation, shRNA, Control