The SETD7–H3K4me1–APLNR Axis Couples Epigenetic Signaling to Orchestrate Endoderm Specification in Human Pluripotent Stem Cells

The differentiation of human pluripotent stem cells into definitive endoderm (DE), a pivotal event in embryonic development, remains incompletely elucidated with respect to its epigenetic control. Here, we identified SETD7, a histone H3K4 monomethyltransferase (H3K4me1), as dynamically upregulated during DE specification, with its expression correlating strongly with the key lineage marker SOX17. Genetic knockdown of SETD7 impaired DE formation, as evidenced by reduced expression of DE markers and disruption of APLNR-mediated PI3K/AKT/mTOR signaling—a phenotypic defect recapitulated by pharmacological inhibition of APLNR. Mechanistically, SETD7 facilitates H3K4me1 deposition at enhancer regions of APLNR, thereby promoting its transcriptional activation. Our findings unveil an epigenetic-regulatory axis wherein SETD7-mediated enhancer priming drives APLNR expression, ultimately activating the PI3K/AKT/mTOR cascade to promote endodermal commitment. Through an integrated approach incorporating doxycycline-inducible SETD7 knockdown in hESCs, pathway-specific pharmacology, ChIP-qPCR, and 3D organoid modeling, we decipher the spatiotemporal regulatory network governed by SETD7 during DE specification. This study highlights the critical crosstalk between histone modification landscapes and signal transduction during cell fate determination and establishes a novel strategy for manipulating differentiation trajectories. Overall design: In this study, we revealed the molecular mechanism by which the SETD7-H3K4me1-APLNR axis regulates DE differentiation. Initially, during the epigenetic initiation phase, SETD7 mediates H3K4me1 modification at APLNR enhancer regions, thereby facilitating APLNR transcriptional activation. Subsequently, the APLNR protein is transported from the cytosol to the plasma membrane, where it becomes properly anchored within the lipid bilayer. Following membrane localization, the APLNR receptor engages with its cognate ligand Apelin, resulting in receptor activation and subsequent initiation of downstream PI3K/AKT/mTOR signaling cascade. This signal transduction ultimately converges on the transcriptional regulation of endodermal marker genes, orchestrating DE lineage specification.