Resolving human α versus β cell fate allocation for the generation of stem cell-derived islets

Generating stem cell-derived glucagon-producing α (SC-α cells) and insulin-producing β cells (SC-β cells) allows to engineer an in vitro biomimetic of the islet of Langerhans, the micro-organ controlling blood glucose, however, there is still a major knowledge gap in the mode and mechanism by which human SC-α and β cells are specified. Mouse studies postulated that Aristaless Related homeobox (Arx) and Paired box 4 (Pax4) transcription factors cross-inhibit each other in endocrine progenitors to promote α or β cell fate allocation, respectively. To test this model in human, we generated an ARXCFP/CFP; PAX4mCherry/mCherry double knock-in reporter induced pluripotent stem cell (iPSC) line to combine time-resolved cell lineage labeling with high-resolution single cell multiomic analysis. Strikingly, lineage labelling and tracing, proteomic and gene regulatory network (GRN) analysis and potency assays revealed a human specific mode and regulatory logic of α versus β cell fate allocation. Importantly, pharmacological perturbation using drugs previously proposed to trigger α-to-β cell transdifferentiation or identified via our GRN analysis led to enhanced endocrine induction and directed α vs β cell fate commitment. Thus, shedding light on basic mechanisms of endocrine induction and fate segregation not only paves the way to engineer islets from pluripotent stem cells, but also has broader implications for cell-replacement therapy, disease modelling and drug screening. ### Competing Interest Statement The authors have declared no competing interest. Source data will be provided upon publication. Any other data supporting the findings of this study are available from the corresponding author on reasonable request.