Fig. 2: MyoD plays a crucial role in inducing MT eRNAs. | Nature Communications

Fig. 2: MyoD plays a crucial role in inducing MT eRNAs.

From: MyoD induced enhancer RNA interacts with hnRNPL to activate target gene transcription during myogenic differentiation

Fig. 2

a Unsupervised clustering of TF binding at eRNA TSSs in MB or MT. The color code indicates the Pearson correlation coefficient (PCC) between two TFs at their binding sites. b Comparison between GRO-seq, total and PolyA+ RNA-seq signals in the MyoD binding proximal regions (±1 kb of the center of MyoD-binding sites) at SEs and TEs. c MyoD ChIP-seq signals within ±2.5 kb flanking TSSs of up-regulated (left) and down-regulated (right) eRNAs. d Distribution of averaged GRO-seq signals from SEs or TEs in WT or MyoD knockout (MyoD−/−) cells. e Illustration of eRNA down-regulation in MyoD−/− vs WT cells by GRO-seq tag counts on seRNA-1, -2, and -5. The bar graph shows the quantification of GRO-seq signals in RPKM. f qRT-PCR measurement of seRNAs in the differentiating MyoD−/− vs WT cells. g qRT-PCR detection of seRNAs from 48-h-differentiated C2C12 cells transfected with either control or MyoD siRNA. h Top: 10T1/2 cells were transfected with either control or MyoD expressing plasmid; the cells were collected in growth medium (GM) or differentiate medium for 48 h (DM). The relative expression of seRNAs, MyoD and Myogenin were measured by semi-quantitative RT-PCR. Bottom: Western blot confirmed MyoD overexpression. i MyoD ChIP-PCR at the TSS of seRNA-1 or seRNA-2 in MT cells. j Luciferase reporter activity of seRNA promoter was detected in 48-hr-differentiated C2C12 cells transfected with either control or MyoD siRNA. k Luciferase reporter activity of the above seRNA promoter in 10T1/2 cells overexpressing MyoD. l Distribution of MyoD ChIP-seq signals on the TSSs of seRNA-1, -2, and -5 in 10T1/2 cells overexpressing MyoD. Data in f, g, i represent the average of three independent experiments ± s.d. Data in b are presented in boxplot. Center line, median; box limits, upper and lower quartiles; whiskers, 1.5x interquartile range. Statistical analysis was performed by two-way ANOVA with Sidak’s post-hoc test (f) or two-tailed unpaired Student’s t-test (g, i, j, k), n.s., not significant, *P < 0.05, **P < 0.01 and ***P < 0.001. Source data are provided as a Source Data file.

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