Fig. 2

Shifting the PRE position to generate a new epigenetic signature. a Schematic representation of the dac TAD and its upstream TAD in WT, ΔPRE1 flies, and PRE1_UP flies. b CUT&RUN profiles for H3 K27 me3 and PH at the dac gene locus in WT, ΔPRE1, PRE1_UP flies. PRE1, PRE2, and the upstream PRE insertion site are indicated by gray bars. Note that In the PRE1_UP line, H3 K27 me3 and PH profiles were aligned to a custom genome in which PRE1 was removed from its endogenous location and inserted at the ectopic site (see the “Methods” section). RNA-seq track of late embryos is shown (modENCODE). c Hi-C score (see the “Methods” section) maps of a 300 kb region at 3 kb resolution on chromosome 2L, including the dac gene locus in WT, ΔPRE1 and PRE1_UP third instar imaginal leg disc. Violet bars indicate the positions of the PREs. Gray bars indicate the position of the TAD boundary. Black arrows indicate gene promoters of the dac and the CG5888. The black dashed rectangle indicates higher contact regions of chromatin marked by H3 K27 me3. d Insulation profile shown at 3 kb resolution along the dac 200 kb region in larval WT and ΔPRE1 (top panel, green) and PRE1_UP (bottom panel, light blue) mutant fly lines is shown as the mean value (line) ± the standard deviation (shaded area) over the insulation scores (IS) computed using 5 different values of the window parameter (w = 100, 150, 200, 250, and 300 kb, see the “Methods” section). eP-values from the comparisons of insulation scores (IS) at the left and right TAD borders between the WT and the corresponding fly line (see Methods). The p-values resulted from a two-sided Welch t-test between the WT condition and each of the mutants at the corresponding locus. f Quantification of dac PRE loop (left) Hi-C interaction score in WT, ΔPRE1, and PRE1_UP mutant flies. The left panel shows quantification of the endogenous PRE loop WT, ΔPRE1, and PRE1_UP; the right panel quantifies the PRE loop between endogenous PRE2 and upstream PRE1. Reported p-values result from the comparisons between WT, ΔPRE1, and PRE1_UP mutant distributions using the unpaired two-sided Wilcoxon statistical test. The number of points per boxplot is reported. Boxplots show median (central line), Q1 = 25 th and Q3 = 75 th percentiles (box limits), and Q1 + 1.5 × IQR to Q3 + 1.5 × IQR (whiskers), where IQR is the interquartile range. g Differential Hi-C interaction score maps (PRE1_UP mutant vs ΔPRE1) of a 300 kb region of the dac gene in third instar imaginal leg disc (see the “Methods” section). Red regions indicate higher contacts in PRE1_UP mutant, blue regions indicate higher contacts in ΔPRE1. The black circle indicates the position of the PRE loop. Violet bars indicate the positions of the PREs. Black arrows indicate the promoters of the dac and the CG5888 genes. The black dashed rectangle indicates higher contact regions of chromatin marked by H3 K27 me3