Fig. 3: rs6048205-G affected pancreatic differentiation through FOXA2 expression. | Nature Communications

Fig. 3: rs6048205-G affected pancreatic differentiation through FOXA2 expression.

From: A noncoding variant confers pancreatic differentiation defect and contributes to diabetes susceptibility by recruiting RXRA

Fig. 3

a Upper: Schematic diagram showing the genes located within 2 Mb away from the rs6048205 locus. Lower: Heatmap showing the expression of these rs6048205-adjacent genes detected by RNA-seq. b mRNA levels of FOXA2 in DE, PP1, PP2 stage detected by RT-qPCR. Gene expression levels were normalized to GAPDH and then further normalized to the A/A group (n = 4 independent experiments in DE and PP1, n = 3 independent experiments in PP2). c Protein levels of FOXA2 in PP2 stage detected by Western blot (n = 3 independent experiments). GAPDH was used as a loading control. d Schematic diagram showing the construction of dox-inducible FOXA2 overexpression system in AAVS1 locus. e Protein levels of FOXA2, PDX1 and NKX6-1 in PP2 stage detected by Western blot (n = 3 independent experiments). GAPDH was used as a loading control. f Representative immunofluorescence images of PP2 cells stained by NKX6-1 (Green). Scale bar, 100 µm. g Quantitative analysis for (f) and (Supplementary Fig. 5e) (n = 12 images). h Scatterplot showing differentially expressed genes identified by RNA-seq of PP2 cells of FOXA2 with and without dox (n = 2 independent samples). Up-regulated and down-regulated genes upon FOXA2 overexpression were shown in red and blue, respectively. i GO analysis of the up- and down-regulated genes in FOXA2 overexpressed PP2 cells. j Left: Venn diagram showing the overlap of differentially expressed genes (DEGs) in SNP-G affected genes and DEGs upon FOXA2 overexpress; Right: GO analysis of these overlapped genes. k The visualization of FOXA2 binding at NKX6-1 gene locus. A dotted box shows a potentially binding site analyzed from ChIP-seq data (GSE114102). l ChIP-qPCR analysis for FOXA2-binding in PP2 cells at NKX6-1 locus (n = 3 independent experiments). m Firefly luciferase/Renilla luciferase results showing the regulation of FOXA2 on NKX6-1 regulatory elements related to (k) (n = 3 independent experiments). Data are presented as the mean ± SD. Statistical significance was determined using the unpaired, two-tailed t-test in (a, b, g, l, m) (ns not significant), one-sided hypergeometric test with multiple comparison adjustments was used in (i, j). Source data are provided as a Source Data file.

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