Extended Data Fig. 6: Epigenomic evaluation of POU2F3, C11orf53/OCA-T1, and COLCA2/OCA-T2 in SCLC-P cell lines.

(a) Comparison of POU2F3, C11orf53, and HA-COLCA2 ChIP-seq peak overlap in the indicated cell lines. For each protein, peaks represent the ones that are consistently identified from 2-3 replicate of experiments. Detailed information is in method. (b) Annotation of POU2F3 and C11orf53 overlapping peaks NCI-H211 cells. (c) Position weight matrix of discovered motif enriched on POU2F3/C11orf53 co-binding sites by MEME from NCI-H211 cell line. (d) Western blot analysis of lentivirally overexpressed constructs used for ChIP-qPCR and gene complementation assay in NCI-H211 cells (n = 2). (e) Sequential ChIP-qPCR analysis of overexpressed HA-C11orf53 (NCI-H211) or HA-COLCA2 (NCI-H1048) co-expressed with FLAG-POU2F3 binding sites in NCI-H211 and NCI-H1048 cells respectively with anti-FLAG (1^st IP) or protein G beads (1^st IP) and anti-HA (2^nd) antibodies. The enrichment is adjusted to the input amount. Two biological replicates are performed and represented as individual dots; the bar value represents the average enrichment over input of two biological replicates. (f) Western blot analysis of POU2F3 or C11orf53 in NCI-H211 (Cas9) cells transduced with indicated sgRNAs. Samples were collected 4 days post infection (n = 1). (g) RNA-seq analysis comparing mRNA changes following C11orf53, COLCA2, or POU2F3 knockout compared to control in three SCLC-P cells. RNA was collected five- or six-days post sgRNA infection. Each dot represents the log₂fold-change of a single protein-coding genes (read outs > = 10). (h) Western blot analysis of FLAG-POU2F3, HA-C11orf53, and FLAG-GFP expression in murine YT330 SCLC cells. Data is representative of two biological replicates. Uncropped gel is provided in Supplementary Fig. 1i. Source data for gene expression changes upon knockout in different cells are provided in Supplementary Table 2. sgRNA and primer sequences are provided in Supplementary Table 6.