Hypoxia and loss of GCM1 expression prevents differentiation and contact inhibition in human trophoblast stem cells [ATAC-Seq]

The placenta develops alongside the embryo and nurtures fetal development to term. During the first stages of embryonic development, due to low blood circulation, the blood and ambient oxygen supply is normally very low (~1-2% O2) and gradually increases upon placental invasion. While a hypoxic environment is associated with stem cell self-renewal and proliferation, persistent hypoxia may have severe effects on differentiating cells and could be the underlying cause of placental disorders. We find that human trophoblast stem cells (TSC) thrive in low oxygen, whereas differentiation of syncytiotrophoblast (STB) and extravillous trophoblast (EVT) is negatively affected by hypoxic conditions. We find that the pro-differentiation factor GCM1 (human Glial Cell Missing-1) is downregulated in low oxygen, and there is substantial reduction of GCM1-regulated genes in hypoxic conditions. Knock-out of GCM1 in TSC caused impaired EVT and STB formation and function, reduced expression of genes that respond to differentiation, and resulted in maintenance of self-renewal genes. Treatment with a PI3K inhibitor reported to reduce GCM1 protein levels likewise counteracts spontaneous or directed differentiation. Chromatin immunoprecipitation of GCM1 showed enrichment of GCM1-specific binding near key transcription factors upregulated upon differentiation including the contact inhibition factor CDKN1C. Loss of GCM1 resulted in downregulation of CDKN1C and corresponding loss of contact inhibition, implicating GCM1 in regulation of this critical process.