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. 2020 Aug 3;9:e58659. doi: 10.7554/eLife.58659

Figure 1. LncRNA expression and regulation during pancreatic differentiation.

(A) Stages of directed differentiation from human embryonic stem cell (hESCs) to hormone-producing endocrine cells. The color scheme for each stage is used across all figures. (B) K-means clustering of all lncRNAs expressed (RPKM ≥ 1) during pancreatic differentiation based on their expression z-score (mean of n = 2 independent differentiations per stage; from CyT49 hESCs). (C,D) Left: Scatterplots comparing the expression of early (C) and late (D) expressed endodermal transcription factors (TFs) with the expression of their neighboring lncRNAs across 38 tissues. The dot color indicates the germ layer of origin of these tissues. Pearson correlation coefficients and p-values (t-test) are displayed. Right: Distribution of the Pearson correlation coefficients for each TF with all Ensembl 87 genes across the same 38 tissues. Dashed lines denote the correlation for the neighboring lncRNA, which for all lncRNAs shown is higher than expected by chance. See also Figure 1—figure supplement 1 and Figure 1—source data 1.

Figure 1—source data 1. Identification, regulation, and characterization of lncRNAs during pancreatic differentiation.
(A) Gene expression during pancreatic differentiation (RPKM). (B) lncRNA-proximal TFs, by cluster in correlation heatmap (Figure 1—figure supplement 1C). (C) GO enrichment and KEGG pathway analysis for each cluster in the correlation heatmap (Figure 1—figure supplement 1D).

Figure 1.

Figure 1—figure supplement 1. Characterization of lncRNAs expressed during pancreatic differentiation.

Figure 1—figure supplement 1.

(A,B) Left: Expression of the single nearest coding genes (±1000 kb) in cis to transcribed and non-transcribed lncRNAs at the DE stage (A) or PP2 stage (B). Log2 transformed mean expression values (RPKM + pseudocount) from two biological replicates were used to generate the box plots (****, p-value<0.0001, Wilcoxon rank sum test). Right: Corresponding cumulative distance distribution functions. (C) Heatmap of the hierarchically clustered expression correlations (Spearman’s rho) of all RNAs transcribed during pancreatic differentiation (with RPKM ≥ 1 in at least ten out of 38 tissues). Transcription factor (TF)-encoding mRNAs, lncRNAs (all), dynamically expressed lncRNAs (RPKM ≤ 1 in at least one stage (ESC to PP2)), and TF-proximal lncRNAs are highlighted above the heatmap. Clusters 8 and 10 are significantly enriched for all of these RNAs (*, p-value<0.03, Fisher test). (D) Gene ontology and KEGG pathway analysis for all coding genes in cluster 8 (p-value<0.05, Fisher test). The full list of significantly enriched terms is shown in Figure 1—source data 1C. (E–H) H3K4me3 and H3K27me3 ChIP-seq tracks of loci containing lncRNAs GATA6-AS1 (A), LINC00261 (B), PDX1-AS1/PLUTO (C), or SOX9-AS1 (D) during pancreatic differentiation of CyT49 hESCs.