iPSC-Derived Pancreatic Progenitors Lacking FOXA2 Reveal Alterations in miRNA Expression Targeting Key Pancreatic Genes

Recently, we reported that forkhead box A2 (FOXA2) is required for the development of human pancreatic alpha- and beta-cells. However, whether miRNAs play a role in regulating pancreatic genes during pancreatic development in the absence of FOXA2 expression is largely unknown. Here, we aimed to capture the dysregulated miRNAs and to identify their pancreatic-specific gene targets in pancreatic progenitors (PPs) derived from wild-type induced pluripotent stem cells (WT-iPSCs) and from iPSCs lacking FOXA2 (FOXA2(-/-)iPSCs). To identify differentially expressed miRNAs (DEmiRs), and genes (DEGs), two different FOXA2(-/-)iPSC lines were differentiated into PPs. FOXA2(-/-) PPs showed a significant reduction in the expression of the main PP transcription factors (TFs) in comparison to WT-PPs. RNA sequencing analysis demonstrated significant reduction in the mRNA expression of genes involved in the development and function of exocrine and endocrine pancreas. Furthermore, miRNA profiling identified 107 downregulated and 111 upregulated DEmiRs in FOXA2(-/-) PPs compared to WT-PPs. Target prediction analysis between DEmiRs and DEGs identified 92 upregulated miRNAs, predicted to target 1498 downregulated genes in FOXA2(-/-) PPs. Several important pancreatic TFs essential for pancreatic development were targeted by multiple DEmiRs. Selected DEmiRs and DEGs were further validated using RT-qPCR. Our findings revealed that FOXA2 expression is crucial for pancreatic development through regulating the expression of pancreatic endocrine and exocrine genes targeted by a set of miRNAs at the pancreatic progenitor stage. These data provide novel insights of the effect of FOXA2 deficiency on miRNA-mRNA regulatory networks controlling pancreatic development and differentiation.

Automated summary

Recently, the importance of forkhead box A2 (FOXA2) in pancreatic cell development has been reported. However, the role of miRNAs in regulating pancreatic genes in the absence of FOXA2 expression is not well understood. This study aimed to identify dysregulated miRNAs and their targets in pancreatic progenitors (PPs) derived from induced pluripotent stem cells (iPSCs) with and without FOXA2 expression. The results showed that FOXA2 deficiency led to reduced expression of PP transcription factors and genes involved in pancreatic development and function. Furthermore, miRNA profiling revealed dysregulated miRNAs in FOXA2-deficient PPs, with predicted targets in downregulated genes. Overall, this study provides insights into the regulatory networks controlling pancreatic development and differentiation in the absence of FOXA2 expression.