Immediate early response 3 interacting-protein 1 (IER3IP1) is an endoplasmic reticulum (ER) resident protein, highly expressed in pancreatic cells and the developing brain cortex. Homozygous mutations in IER3IP1 have been found in individuals with microcephaly and neonatal diabetes, yet the underlying mechanism causing β-cell failure remains unclear. Here, we used differentiation of genome-edited stem cells into pancreatic islet cells to elucidate the molecular basis of IER3IP1 neonatal diabetes. Using clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated protein 9 (Cas9), we generated two distinct IER3IP1 mutant human embryonic stem cell lines: a homozygous knock-in model of a patient mutation (IER3IP1V21G), and a knockout (KO) model (IER3IP1−/−). While these mutant stem cell lines differentiated normally into definitive endoderm and pancreatic progenitors, we observed that IER3IP1-KO stem cell-derived islets (SC-islets) presented a significant decrease in β-cell numbers and elevated ER stress. Retention using selective hooks assay revealed a threefold reduction in ER-to-Golgi trafficking of proinsulin in IER3IP1 mutant β-cells. Additionally, IER3IP1 mutant SC-islets implanted into immunocompromised mice displayed defective human insulin secretion, indicating the deleterious impact of IER3IP1 mutations on β-cell function. Our study provides valuable insights into the role of IER3IP1 in human β-cell biology and establishes a useful model to investigate ER-to-Golgi trafficking defects within β-cells.
IER3IP1 mutations are linked to the development of microcephaly, epilepsy, and early-onset diabetes syndrome 1. However, the underlying molecular mechanisms of cell dysfunction are unknown.
Using targeted genome-editing, we generated specific IER3IP1 mutations in human embryonic stem cell lines that were differentiated into pancreatic islet lineages.
Loss of IER3IP1 resulted in a threefold reduction in endoplasmic reticulum-to-Golgi trafficking of proinsulin in stem cell-derived β-cells, leading to β-cell dysfunction both in vitro and in vivo.
Loss of IER3IP1 also triggered increased markers of endoplasmic reticulum stress, indicating the pivotal role of the endoplasmic reticulum-to-Golgi trafficking pathway for β-cell homeostasis and function.
Time-lapse of the RUSH assay in WT β-cell starting 6 min after biotin addition, with a picture acquired every 2 min. Scale bar = 4 µm.
Time-lapse of the RUSH assay in WT β-cell starting 6 min after biotin addition, with a picture acquired every 2 min. Scale bar = 4 µm.
Time-lapse of the RUSH assay in IER3IP1−/− β-cell starting 6 min after biotin addition, with a picture acquired every 2 min. Scale bar = 5 µm.
Time-lapse of the RUSH assay in IER3IP1−/− β-cell starting 6 min after biotin addition, with a picture acquired every 2 min. Scale bar = 5 µm.
This article contains supplementary material online at https://doi.org/10.2337/figshare.27245415.
