GFI1 occupies the ELDR promoter and regulates ELDR expression. (A) ChIP-seq data from THP-1 cells (GSE63222²³) revealing enrichment of GFI1 recruitment at the 5′ end of the ELDR locus. (B) Schema of the human ELDR locus, which encompasses a region of 19 670 bases and encodes a lncRNA of 2941 bases. The ELDR locus is located downstream of the EGFR gene on human chromosome 7. (C) ChIP-seq data from THP-1 cells (GSE63222) reveal that GFI1 and LSD1 are evicted from the ELDR promoter after treatment with the LSD1i OG86. (D) ChIP–quantitative polymerase chain reaction (qPCR) with THP-1 cell extracts and an anti-GFI1 antibody using primers for the ELDR 5′ region. The GFI1 promoter itself served as a positive control and the ERICD locus as a negative control (n = 3 biological replicates). (E) Induction of ELDR expression after treatment of THP-1 cells with the LSD1i GSK2879552 (n = 3 biological replicates). (F) Reduction of GFI1 expression by short hairpin RNA (shRNA) (shGFI1) as compared with control nontargeting shRNA (shCtr) leads to an increase of ELDR expression in THP-1 cells. Western blot revealing knockdown of GFI1 in THP-1 cells. (G) Effect of the induction of endogenous ELDR expression by LSD1i on nearby genes within ±200 kbp. THP-1 cells were treated with LSD1i (GSK2879552) or DMSO (control). Reverse transcription–qPCR (RT-qPCR) measurement of EGFR, VOPP1, and LANCL2 RNA expression (n = 2 biological replicates for the PCR). (H) Overexpression of Gfi1 (Gfi1o/e) in RAW 264.7 cells as compared with NT cells leads to Eldr repression (n = 3 biological replicates). Western blot revealing overexpression of GFI1 in RAW 264.7 cells. (I) RT-qPCR analysis reveals overexpression of ELDR in THP-1 stable cell lines established with a lentiviral vector directing the expression of ELDR (ELDR-C1, ELDR-C3, ELDR-C5) or with an empty vector (EV1, EV2) (n = 3 biological replicates for the PCR). (J) RT-qPCR analysis reveals overexpression of ELDR in Mono-Mac-1 stable cell lines established with a lentiviral vector directing the expression of ELDR (ELDR-C5 and ELDR-C7) or with an empty vector (EV1, EV2) (n = 3 biological replicates for the PCR). (K) Effect of ectopic overexpression of ELDR in established ELDR-overexpressing cell lines on nearby genes within ±200 kbp. RT-qPCR measurement of EGFR, VOPP1, and LANCL2 RNA expression (n = 3 biological replicates for the PCR). (L) Expression of ELDR via RT-qPCR in different subtypes of AML cell lines (n = 3 technical replicates for the PCR). Data are presented as mean + standard deviation (SD). ∗∗P < .01; ∗∗∗P < .001; ∗∗∗∗P < .0001. P values were calculated using parametric unpaired t test (panels D-F,H,K); 1-way analysis of variance (ANOVA) with Holm-Šídák's for multiple comparisons test (panels G,I-J,L). DMSO, dimethyl sulfoxide; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; ns, not significant.