Fig. 2: VNTR6-1 affects the TERT-FL:TERT-β splicing ratio.

a G4-ChIP results within the TERT region in the cell lines HEK-293T (VNTR6-1: Long/Long) and NA18507 (YRI, 1000 G, VNTR6-1: Short/Short) display mismatches (%) during DNA synthesis, reflecting polymerase stalling after G stabilization in both the plus (blue) and minus (orange, direction of TERT transcription) genome strands. The genomic region of TERT intron 6 includes VNTR6-1 (24-66.5 copies of the 38-bp repeat unit), VNTR6-2, G4 in the minus strand (polymorphic G4 within VNTR6-1 and constitutive upstream G4), and CRISPR/Cas9 guide RNAs for excising VNTR6-1. The sequence logo shows the consensus 38-bp VNTR6-1 repeat unit in UMUC3 cells based on long-read WGS. b Agarose gels of RT‒PCR products amplified from the cDNA of corresponding samples; gDNA–genomic DNA negative control; HPRT1 -endogenous normalization control. e Densitometry results of the PCR amplicons in the plot (b). The differences in the TERT isoform ratios are further explored in Supplementary Fig. 11. Experiments in UMUC3 cells comparing TERT splicing and isoform-specific expression after 72 h of treatment with G4 stabilizing ligands, normalized to HPRT1 in the WT (c, f) and V6.1-KO (d, g) cell lines. c, d A representative agarose gel of SYBR-Green RT‒qPCR products detecting several isoforms with primers located in exons 6 and 9. The extra PCR band, marked by a red arrow in panels (c and d), is further explored in Supplementary Fig. 12. f, g Densitometry analysis of the corresponding agarose gels evaluating TERT-FL (%) relative to the total PCR products. All analyses are based on three independent experiments, presented with means ± SD. One representative gel per experiment is shown. Comparisons were made against the vehicle control (DMSO). P-values are for unpaired two-sided Student’s T test. The source data are provided in the Source Data file.