The cellular response of immortalized TRF2dB fibroblast cells by 4-hydroxytamoxifen drug treatments

In our study, we have successfully generated a cell line that stably expresses a mutant TRF2 with a truncated N-terminal basic domain from BJ normal human fibroblasts. This mutant named, TRF2∆B, can artificially loop out telomeric DNA as ECTR in circular forms, mediated by XRCC3 holiday junction resolution protein and SLX4 associated recombination endonucleases. To avoid the deleterious effect on cell growth resulting from constant TRF2∆B expression, we fused TRF2ΔB with an estrogen receptor ligand-binding domain variant ERT2 to keep TRF2∆B inactive in the BJTL cells, and in the presence of 4-hydroxytamoxifen (4-OHT),which binds to ERT2, TRF2∆B becomes active. With this system, we aim to investigate the cellular response of immortalized BJ fibroblast cells to 4-OHT drug treatments. Cellular response in immortalized human fibroblast transduced with ERT2-TRF2dB gene was measured at 0, 12, 24, 36, 48 and 60 hours of 1 µM 4-hydroxytamoxifen (4-OHT). IFNb gene expression in immortalized human fibroblast transduced with vector was measured at 0 and 48 hrs of 1 µM 4-hydroxytamoxifen (4-OHT) as controls. Cellular expression of treated ERT2-TRF2dB cells are normalized to untreated ERT2-TRF2dB cells and compared to the expressions of 4-OHT treated vector fibroblast cells that are normalized to untreated vector fibroblast cells. Three independent experiments were performed at each time for each experiment. Liuh-Yow, Chen