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

Stem cell-derived glucagon-(α) and insulin-producing (β) cells allow to engineer in vitro biomimetics of islet of Langerhans, the micro-organ controlling glycemia; however, a knowledge gap in the mechanism by which human stem cell-derived α and β cells are specified persists. 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/β fate allocation, respectively. To test this model in human, we combined lineage labeling with single-cell multiomic analysis in our newly generated ARXCFP/CFP; PAX4mCherry/mCherry knock-in induced pluripotent stem cell reporter line. Lineage tracing, proteomic and gene regulatory network analysis and potency assays revealed a human specific regulation of α/β cell fate allocation. Pharmacological perturbations previously proposed to trigger α-to-β transdifferentiation or identified by our gene regulatory network led to enhanced endocrine induction and directed α/β cell fate. Studying mechanisms of endocrinogenesis and fate segregation enables to engineer islets in vitro, and has broader implications for cell-replacement therapy, disease modelling and drug screening.