Fig. 8: HNF1A H126D mutation reduced the ability of HNF1A to upregulate GLUT2 gene expression. | Nature Communications

Fig. 8: HNF1A H126D mutation reduced the ability of HNF1A to upregulate GLUT2 gene expression.

From: Decreased GLUT2 and glucose uptake contribute to insulin secretion defects in MODY3/HNF1A hiPSC-derived mutant β cells

Fig. 8

a RT-qPCR analysis of GLUT2, GLUT1 and GLUT3 transcripts in AD-293 cells overexpressed with GFP (green) and various WT (red), H126D (blue) and P291fsinsC (purple) HNF1A constructs. n = 3 independent experiments; P-value for GLUT2 = 0.0156 (WT), 0.0080 (H126D), 0.0227 (P291fsinsC); GLUT1 = 0.0296 (H126D); GLUT3 = 0.006 (WT), 0.0063 (H126D), 0.0141 (P291fsinsC). b Immunohistochemistry stain for GLUT2 (red) and nuclear stain using DAPI (blue) in AD-293 cells overexpressed with GFP and various HNF1A constructs. n = 3 independent experiments. (scale bar:100 μm). c Immunofluorescence data was quantified by cell counting using ImageJ across three representative images per condition in AD-293 cells overexpressed with GFP (green) and various WT (red), H126D (blue) and P291fsinsC (purple) HNF1A constructs. d Luciferase reporter analysis of GLUT2 promoter activity in AD-293 cells overexpressed with GFP (green) and various WT (red), H126D (blue) and P291fsinsC (purple) HNF1A constructs (n = 3). e RT-qPCR analysis of GLUT2 transcripts in EndoC-βH1 cells transfected with HNF1A siRNA (si-HNF1A) and non-targeting siRNA as negative control (si-NT). n = 6 independent experiments. p = 0.0012. f ChIP-qPCR analysis of HNF1A binding onto GLUT2 promoter in hPSC-derived endocrine progenitors, using HNF1A antibody (red) and negative control IgG (blue). n = 3 independent experiments. One-way ANOVA was performed for group comparison (p = 0.0132, F-critical = 6.3419). Tukey post-hoc test was done for pairwise comparison, *p = 0.0209, **p = 0.0152. g Summary diagram showing the components of stimulus-secretion coupling in human β cell affected by HNF1A H126D mutation. WT wild-type, P1 patient 1, P2 patient 2. GFP green fluorescent protein. ChIP-Seq Chromatin immunoprecipitation sequencing. RT-qPCR quantitative reverse transcription polymerase chain reaction. hPSC human pluripotent stem cells. For all statistical analysis: Error bars represent standard error of mean (SEM). Unpaired one-tailed Student’s t-test was performed for all statistical analysis, otherwise stated. Asterisk indicates P-value < 0.05. n.s. non-significant. “See also Figs. S7 and S11.” Source data are provided as a Source data file.

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