Regulatory mechanisms of neural stem cell proliferation by hypoxia-inducible factor

Neural stem cells (NSCs) reside in vivo in hypoxic environments, and NSC proliferation is enhanced in vitro under hypoxic conditions.Various adaptive responses to hypoxia are mediated by hypoxia-inducible factors (HIFs), a family of basic helix-loop-helix Per-Arnt-Sim (PAS) transcription factors.Necdin, a MAGE (melanoma antigen) family protein, is expressed abundantly in postmitotic neurons and possesses potent anti-mitotic and anti-apoptotic activities.We here report that hypoxia induces degradation of the necdin protein in primary NSCs by HIF-mediated ubiquitin-proteasome system.Necdin was expressed in primary NSCs prepared from the ganglionic eminences of mouse embryos.Hypoxia enhanced neurosphere formation of NSCs, in which the necdin protein level was significantly reduced.Primary NSCs prepared from necdin-deficient mice exhibited higher rates of proliferation and apoptosis than those from wild-type mice in normoxia, whereas there were no significant differences in the proliferation and apoptosis rates between necdin-deficient and wild-type NSCs in hypoxia.HIF-2α was predominantly expressed in hypoxic NSCs, where expression of HIF-responsive genes was upregulated.HIF-2α interacted with necdin via its PAS domain, which enhanced necdin ubiquitination.Lentivirus-mediated expression of the PAS domain in primary NSCs promoted necdin degradation and enhanced NSC proliferation in normoxia, whereas a small-molecule inhibitor of HIF-2α translation stabilized the necdin protein and reduced NSC proliferation in hypoxia.These results suggest that oxygen tension regulates the necdin protein level in NSCs through HIF-2α-mediated proteasomal degradation to modulate their proliferation and apoptosis.