Fig. 2
MYC represses BCL2 expression. a GI50 values of MYC (n = 4; GLC1, H82, H524, GLC2), MYCN (n = 4; GLC8, H69, SBC4, COR-L303), and MYCL (n = 3; H1092, H2029, SBC7) human small cell lung cancer (SCLC) cell lines treated with navitoclax for 72 h (n = 3). b GI50 values of Myc paralog-activated CRISPRa cells treated with navitoclax for 96 h (n = 3). c Relative mRNA levels (quantitative reverse transcription polymerase chain reaction (qRT-PCR)) of Myc and Bcl2 in Mycn-activated CRISPRa cells with Tet-inducible Myc knockdown by shRNA at 72 h after doxycycline treatment (n = 3). Data were normalized to 18S rRNA. d Viability screening of Mycn-activated CRISPRa cells after Myc knockdown treated with navitoclax for 96 h (n = 3). e GI50 values of viability screening in d (n = 3). f DNA methylation at the BCL2 promoter (indicated region) measured by bisulfite sequencing. Lollipop diagram representing methylated (black) and unmethylated (white) CpGs in MYC (H82: n = 8, GLC1: n = 8, GLC2: n = 7), MYCN (H69: n = 8, SBC4: n = 8), and MYCL (H1092: n = 7) amplified human SCLC cell lines. g Percentage of methylated CpG residues in MYC (H82, GLC1, GLC2), MYCN (H69, SBC4), and MYCL (H1092) amplified human SCLC cell lines (top). CpG methylation percentage (f) in SCLC cell lines (n = 6) grouped by MYC amplification status (MYC-high n = 3, MYC-low n = 3; bottom). h, i Occupancy at the BCL2 promoter of MYC, MIZ1, and DNMT3a in MYC-amplified cells (GLC1, H82) (h) and of MYCN, MIZ1, and DNMT3a in MYCN-amplified cells (SBC4, H69) (i) measured by chromatin immunoprecipitation (ChIP) quantitative real-time PCR (n = 3). ChIP signal is displayed as percentage of input. IgG (non-specific antibody control) signal was subtracted from ChIP signal of specific antibodies. j Western blot showing BCL2 in MYC-amplified GLC1 cells treated with 1 µM 5-azacytidine for the indicated times. HSP90 was used as loading control. k Relative mRNA expression (qRT-PCR) of BCL2 and DNMT3a in MYC-amplified GLC1 cells treated with control small interfering RNA (siRNA) or DNMT3a siRNA (n = 3). Data were normalized to 18S rRNA. Error bars indicate mean ± SEM. Two-tailed unpaired t tests, ***p < 0.001, **p < 0.01, *p < 0.05. Source data are provided as a Source Data file
