Fig. 2

WRN binds to G4 regions of SHOX promoter to promote SHOX expression. a Illustration of SHOX genomic track showing the promoters cloned for luciferase assay in Fig. 1d, g and G4hunter predicted SHOX G4 regions based on slot blot assay in Fig. 1c and Additional file 1: Fig. 1. b SHOX promoter G4s, MYC G4 (positive control) and random oligo (negative control) sequences subjected to slot blot assay in Fig. 1c. c Slot blot assay of 50 bp ssDNA encompassing G4Hunter predicted G4 regions of SHOX promoters. Oligos were heated and gradually cooled to form G4 structures in vitro, followed by nitrocellulose binding and BG4 antibody staining. d Luciferase assay of SHOX promoters in WRN^WT 293T. e ChIP-qPCR analysis of FLAG-WRN^K577M occupancy in WRN^KO 293 T cells using FLAG antibody for ChIP and PCR primers for amplifying SHOX P2 promoter G4 regions. Negative control is an intergenic region in Chr6. Positive controls (KRAS and αSAT) are known binding targets of WRN. N = 3. t-test (2-tailed, unequal variance) confidence comparing to G4- intergenic region: *p < 0.05, **p < 0.005, ****p < 0.00005. f Luciferase assay of SHOX promoters in WRN^WT and WRN^KO 293T cells rescued with different WRN mutant proteins. g Luciferase assay of SHOX promoter2 (P2) and the different mutated promoters in WRN^WT 293T. h Luciferase assay of mutated SHOX P2 promoters in WRN^WT and WRN^KO 293T cells rescued with different WRN mutant proteins