An oxygen-insensitive Hif-3α isoform inhibits Wnt signaling by destabilizing the nuclear β-catenin complex

Abstract

Hypoxia-inducible factors (HIFs), while best known for their roles in the hypoxic response, have oxygen-independent roles in early development with poorly defined mechanisms. Here, we report a novel Hif-3α variant, Hif-3α2, in zebrafish. Hif-3α2 lacks the bHLH, PAS, PAC, and ODD domains, and is expressed in embryonic and adult tissues independently of oxygen availability. Hif-3α2 is a nuclear protein with significant hypoxia response element (HRE)-dependent transcriptional activity. Hif-3α2 overexpression not only decreases embryonic growth and developmental timing but also causes left-right asymmetry defects. Genetic deletion of Hif-3α2 by CRISPR/Cas9 genome editing increases, while Hif-3α2 overexpression decreases, Wnt/β-catenin signaling. This action is independent of its HRE-dependent transcriptional activity. Mechanistically, Hif-3α2 binds to β-catenin and destabilizes the nuclear β-catenin complex. This mechanism is distinct from GSK3β-mediated β-catenin degradation and is conserved in humans. These findings provide new insights into the oxygen-independent actions of HIFs and uncover a novel mechanism regulating Wnt/β-catenin signaling.Proteins known as hypoxia-inducible factors (HIFs) are important in animals when the amount of oxygen in the air or water drops. These proteins switch on genes that help cells and tissues adapt to the shortage in oxygen, for example, by stimulating the production of red blood cells. Each HIF is made up of two subunits called α and β that only bind to each other when the oxygen levels drop. This two-subunit complex, or 'dimer', then activates a set of genes by binding to a stretch of DNA known as the hypoxia response element. HIFs also play important roles in many different stages of animal development. There are many different HIF proteins that are each present at different developmental stages; this has made them difficult to study.Zhang et al. have found a new form of HIF-3α in zebrafish – called Hif-3α2. The experiments show that this α subunit is not regulated by oxygen, but may still be able to activate genes that have the hypoxia response element. When Hif-3α2 was injected into zebrafish embryos, the body pattern that is normally set up in embryogenesis was disrupted. Further experiments revealed that Hif-3α2 regulates embryo development by destabilizing a protein called β-catenin. This inhibits a cell communication system called Wnt/β-catenin signaling. Zhang et al. also show that the two distinct activities of Hif-3α2 – binding to the hypoxia response element and destabilizing β-catenin – involve two different regions of the protein.Together, Zhang et al.’s findings show that zebrafish Hif-3α2 combines some conventional features of HIF proteins with a unique developmental role. It is likely that human Hif-3α may also work in a similar way, so future studies will focus on understanding the molecular mechanisms responsible for these distinct roles.