Figure 8.

Dual luciferase reporter reveals faster silencing kinetics during differentiation in ΔTelAP1 cells. (A) Illustration of the dual luciferase reporter. The active ES of WT and ΔTelAP1 cells was doubly marked with an Rluc reporter gene downstream of the ES promoter and an Fluc reporter gene upstream of VSG gene. Graphic is not to scale. (B) Analysis of luciferase activity at the ES promoter (Pol I) and at the telomere during differentiation of WT and ΔTelAP1 reporter cell lines. Differentiation of reporter cell lines (n = 3) was induced and luciferase activity was measured at the time points indicated. Luciferase activity is shown as relative light units (RLU) and standard deviation. Time point 0 h was set as 100%. (C) Reintroduction of TelAP1 in ΔTelAP1 reporter cell line slows down ES silencing kinetics during differentiation. Analysis of luciferase activity at the ES promoter and at the telomere during differentiation of ΔTelAP1R reporter cells with and without tetracycline induction. One clone was analyzed in triplicates. WT values of the experiment shown in B were included into the graphs for better comparison. Reintroduction of TelAP1 leads to ES silencing kinetics similar to WT cells. Statistical significance was determined by an unpaired t-test *P < 0.05, **P < 0.01, ***P < 0.001.