Search for author "Tuo Zhang"

Figure 1. HDAC inhibition disrupts p65-mediated repression of Bnip3 gene transcription. Panel A, Postnatal ventricular myocytes were transfected with a Bnip3 luciferase reporter with and without a p65 NF-κB expression vector in the presence and absence of trichostatin A (TSA, 10 nM). Panel B, IKKβ-mediated NF-κB activation represses Bnip3 gene transcription in ventricular myocytes. Postnatal ventricular myocytes were infected with adenoviruses encoding IKKβ (IKKβ) in the presence and absence TSA (10 nM). Endogenous Bnip3 in ventricular myocytes was examined by semiquantitative RT-PCR. Data were normalized to house keeping control gene L32. Control cells (CNTL) were transfected with the eukaryotic expression vector pcDNA3 (panel A) or infected with control adenovirus (panel B). Data are expressed as mean ± S.E. (P<0.05). Experiments were repeated at least n=6 with independent culture conditions using replicates of n=3 for each condition tested, *=statistically different from CNTL; ‡=statistically different from p65; N.S.=not significant.Show Less

Figure 3. Regulation of Bnip3 gene transcription by HDAC1. Panel A, BNIP3 gene transcription in the presence of WT HDAC1 and catalytically inactive HDAC1 mutant (HDAC1H141A). Panel B, Regulation of Bnip3 gene transcription by p65 NF-κB in the presence and absence of WT HDAC1 and mutant HDAC1H141A. Data are expressed as mean ± S.E. (P<0.05). Experiments were repeated at least n=6 with independent culture conditions using replicates of n=3 for each condition tested, *=statistically different from CNTL; †=statistically different from HDAC1; # = statistically different from p65+HDAC1.Show Less

Figure 5. NF-κB elements are required for repression of BNIP3 expression. Schematic of the BNIP3 promoter depicting the NF-κB response elements (NRE) base pairs (-1075/-1069). Postnatal ventricular myocytes were transfected with luciferase reporter plasmids containing WTBnip3 promoter (BNIP3 WT) or mutant Bnip3 promoter, in which the NF-κB elements were deleted and designated (Bnip3 mt), in the presence and absence of p65NF-κB and HDAC1. Data are expressed as mean ± S.E. fold decrease from their respective controls (BNIP3 WT or Bnip3 mt). Experiments were repeated at least n=4 with independent culture conditions using replicates of n=3 for each condition, *=statistically different from Bnip3 wt +p65NFκB; ‡=statistically different from Bnip3WT +HDAC1. Legend; ▪=Bnip3 WT; □=Bnip3mt.Show Less

Figure 7. HDAC1 activity and Bnip3 expression during hypoxia. Panel A, Normoxic control cells (CNTL) or cells subjected to hypoxia (HYPX) in the presence and absence of TSA (10 nM) were assessed for HDAC activity. HDAC1 activity was monitored by following the deacetylation of recombinant fluormetric HDAC substrate HOS341 and is expressed as μmol/min/μg protein; control cells=1.01×10−6 μmol/min/μg protein; cells treated with TSA=6.42×10−7 μmol/min/μg protein; cells subjected to hypoxia=7.91×10−7 μmol/min/μg protein; cells treated with TSA and subjected to hypoxia=5.70×10−7μmol/min/μg protein. Data are presented as percent change from control using n=3 independent experiments and replicates of at least n=3 for each condition tested. Panel B, Bnip3 transcription during hypoxia; Bnip3 gene transcription is increased by 5.0-fold (P<0.001) in cells subjected to hypoxia compared with normoxic control cells; hypoxia-induced Bnip3 gene activation is repressed by WT HDAC1 but not the catalytically inactive mutant HDACH141A; Panels A and B, *=statistically different from normoxic control cells (CNTL); ‡=statistically different from HYPX, N.S.=not significant. Panel C, The NF-κB element is required for suppression of Bnip3 transcription by HDAC1 during hypoxia; Postnatal ventricular myocytes were transfected with luciferase reporter plasmids containing WT BNIP3 promoter (BNIP3 WT) or mutant Bnip3 promoter in which the NF-κB site was deleted and designated (Bnip3 mt) as detailed in Figure 4; *=statistically different from Bnip3 WT normoxic control; ‡=statistically different Bnip3 WT HYPX; †=statistically different from Bnip3 WT HYPX+HDAC1; #=statistically different from Bnip3 mt normoxic control. Legend ▪=Bnip3 WT; □=Bnip3 mt.Show Less