Mathematical Model of a Telomerase Transcriptional Regulatory Network Developed by Cell-Based Screening: Analysis of Inhibitor Effects and Telomerase Expression Mechanisms
Figure 6
Simulation of Ets family transcription factor gain of function at the TERT promoter.
(A), overexpression of ETS2 against the promoter panel. A2780 cells were transfected with the luciferase reporters shown. Each reporter was co-transfected alongside vector control or pCMV-ETS2. 48 h post-transfection, promoter activities were analysed by luciferase assay. Mean ± SEM of three experiments (ns: not significant; *: p<0.05; **: p<0.01). (B), regulation of the ETS2 promoter by the transcription factor panel. A2780 cells were co-transfected with ETS2-luciferase reporter alongside vector control or expression vectors shown. 48 h post-transfection, promoter activities were analysed by luciferase assay. Mean ± SEM of three experiments (ns: not significant; *: p<0.05; **: p<0.01). (C), effect of ETS2 expression on the TERT promoter under MYC inhibition. A2780 cells were co-transfected with the TERT-luciferase reporter and with pCMV control or pCMV-ETS2 with non-targeting or MYC-specific siRNA. 48 h post-transfection, promoter activities were analysed by luciferase assay. Mean ± SEM of three experiments (D), interactions in the ETS2 subnetwork added into the model with cutoffs FC>1.5, p<0.01 from the transfection data. The subnetwork was visualised in Cytoscape [105]. Arrows indicate activation, T-shape indicates repression. (E), Effect of single- and double-node targeting on TERT on-states in the ETS2 modified model. Rule-sets for each node were modified in turn individually (black bars) to simulate constitutive repression or activation. For each rule-set change, statespace was derived and the proportion of system states evolving to attractor states with TERT stably on was quantified. The analysis was repeated in the background of the MYC suppressed rule-set (grey bars).