Extended Data Fig. 5: L1 silencing by CTBP1 restricts distal gene expression.
From: LINE-1 transcription activates long-range gene expression
a, Western blotting showing that CTBP1 KO increases L1 protein expression in K562. b, FACS showing CTBP1 KO increases L1-GFP expression in K562. Center line, mean. n = 2 biological replicates. c, qRT–PCR data showing normalized expression of L1Hs, L1PA15-16, L1PB and L1M subfamilies in control (n = 3) and CTBP1 KO cells (n = 3) with or without reverse transcription (RT). P, two-tailed t test. d, Box plot showing that CTBP1 KO mainly derepresses L1PA1–3 among all repeats in K562. Box plots show median and interquartile range (IQR), and whiskers are 1.5-fold IQR. e, Density plot showing size distribution of the CTBP1-regulated L1s (P < 0.05, DESeq2 analysis) and other L1s in K562. P, two-tailed Kolmogorov–Smirnov test. f, Aggregated line plots showing indicated ChIP enrichments over the CTBP1-regulated L1PAs (P < 0.05, DESeq2 analysis) and all full-length (FL) L1s in K562. g, CTBP1 KO increases expression of genes surrounding the derepressed L1s. Plots show median and interquartile range (IQR) of gene expression change alongside derepressed L1s (black box plots), compared with background (red). h, Bar plot showing the number of full-length L1s detected by H3K27ac ChIP–seq and polyA+ RNA-seq (TPM > 0.05) in K562. i, Zoom-in views of the RNA-seq genome browser tracks at NRN1, its cognate distal L1PA2 and one nearby FARS2 locus, in control and CTBP1 KO K562, independently repeated once with similar results. j, RT–qPCR showing normalized expression of NRN1 and its nearby FARS2 in control (n = 3) and CTBP1 KO cells (n = 3). P, two-tailed t test. n.s., not significant. k,l, Deleting the L1PA2 element, as shown in i and confirmed by the gel image (k), decreases NRN1 expression in NCCIT (l). n = 4 biological replicates × 3 technical replicates. ***P < 0.001; unpaired two-tailed t-test. Error bars, standard deviation (s.d.). m,n, Representative Hi-C interaction matrix and ChIP–seq IGV tracks around NRN1-L1 in wild-type (WT) condition (m) and Sanger sequencing results showing the L1 deletion sites with sgRNA target sequences in red (n).
