The effect of BRD2 knock-out on PC12 cell apoptosis using BRD2-specific siRNA

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Description: Bromodomain 2 (BRD2) is the founding member of the BET family of proteins and is known to be a major histone code interpreter and transcriptional co-regulator. BRD2 is up regulated in neuronal apoptosis, possibly through conserved epigenetic pathways in association with transcription factor E2F and/or trithorax chromatin regulators. The potentially causal role of BRD2 in apoptosis was explored using siRNA gene silencing in undifferentiated neuronal PC12 cells undergoing serum-deprivation induced cell death. Fluorescent microscopy was employed to measure levels of both apoptosis and necrosis after 12 hours of serum starvation using fluorescently labeled Annexin-V, and propidium iodide, respectively. Treatment groups included: untransfected cells, control-siRNA-transfected cells, and Brd2 siRNA transfected cells. A total of four replicates for each treatment group were tested in 2 independent experiments. Chi-square contingency evaluations conducted on intra-well and inter-well variability within treatment groups demonstrated for the most part only insignificant differences. In contrast, inter-treatment chi-square analysis showed significant differences in annexin-V positive cells between Brd2-siRNA transfected cells and controls in the absence of serum. The results indicate that transfection of BRD2 specific siRNA has a diminishing effect on the rate of serum deprivation-triggered apoptosis in undifferentiated neuronal PC12 cells, and preliminarily implicate BRD2 as a necessary regulator of cell death in this paradigm This study thus justifies further investigation into the potential role of BRD2 in naïve as well as neuronal PC12 cell apoptosis ideally through the utilization of stable lines of cells carrying inducible constructs expression BRD2-siRNA.
Language: English
Format: Degree Work