Fig. 3. Chemical dimerization of LSD1 to telomeres is sufficient to enrich TERRA and rescue ALT phenotypes.

a IP-recovered RNA was detected via dot-blot with probes annealing with TERRA or 18S rRNA (refer to Supplementary Fig. 4a for IP efficiency). b RNA pull-down assay was used to evaluate the binding of Streptavidin bead-bound biotinylated (UUAGGG)₈ oligonucleotide to flag-tagged LSD1^FL, LSD1^IDR, LSD1^ΔIDR, LSD1^{ΔIDR, 3KE}, and LSD1^{ΔIDR, K661A} from U2OS cell lysates (n = three independent experiments). c Schematic of LSD1: full-length (LSD1^FL), C-terminus (LSD1^ΔIDR,), N-terminus (LSD1^IDR), mutant for nucleic acid binding (LSD1^{ΔIDR, 3KE}), mutant for demethylation activity (LSD1^{ΔIDR, K661A}). d Dimerization schematic: LSD1 is fused to mCherry and eDHFR, and TRF1 is fused to Halo enzyme and GFP. The dimerizer is TFH, TMP(trimethoprim)- Fluorobenzamide-Halo ligand. e Representative images and f quantification of TERRA (RNA FISH) localization on telomeres after dimerizing overexpressed LSD1^FL, LSD1^IDR, LSD1^ΔIDR, and LSD1^{ΔIDR, 3KE}, and LSD1^{ΔIDR, K661A} to Halo-TRF1 in U2OS cells with dimerizer TFH (n = 108 cells for +TFH alone, n = 87 cells for LSD1^FL, n = 103 cells for LSD1^IDR, n = 113 cells for LSD1^ΔIDR, n = 55 cells for LSD1^{ΔIDR, 3KE}, n = 53 cells for LSD1^{ΔIDR, K661A} over three independent experiments, one-way ANOVA). g Representative images and h quantification of Edu assay showing the rescue effect of newly synthesized telomeric DNA after dimerizing LSD1^FL, LSD1^IDR, LSD1^ΔIDR, LSD1^{ΔIDR, 3KE}, and LSD1^{ΔIDR, K661A} to telomeres in siLSD1 cells (n = 69 cells for siCtr, n = 89 cells for siLSD1, n = 67 cells for LSD1^FL, n = 77 cells for LSD1^IDR, n = 90 cells for LSD1^ΔIDR, n = 75 cells for LSD1^{ΔIDR, 3KE}, n = 81 cells for LSD1^{ΔIDR, K661A} over two independent experiments, one-way ANOVA). Error bars are mean ± SEM. NS not significant. Source data are provided as a Source Data file.