Abstract
The prolyl-3,4-dihydroxylase Ofd1 and nuclear import adaptor Nro1 regulate the hypoxic response in fission yeast by controlling activity of the sterol regulatory element-binding protein transcription factor Sre1. Here, we identify an extra-ribosomal function for uS12/Rps23 central to this regulatory system. Nro1 binds Rps23, and Ofd1 dihydroxylates Rps23 P62 in complex with Nro1. Concurrently, Nro1 imports Rps23 into the nucleus for assembly into 40S ribosomes. Low oxygen inhibits Ofd1 hydroxylase activity and stabilizes the Ofd1-Rps23-Nro1 complex, thereby sequestering Ofd1 from binding Sre1, which is then free to activate hypoxic gene expression. In vitro studies demonstrate that Ofd1 directly binds Rps23, Nro1, and Sre1 through a consensus binding sequence. Interestingly, Rps23 expression modulates Sre1 activity by changing the Rps23 substrate pool available to Ofd1. To date, oxygen is the only known signal to Sre1, but additional nutrient signals may tune the hypoxic response through control of unassembled Rps23 or Ofd1 activity.
Highlights
Eukaryotic cells require oxygen and must adapt to changes in its supply to maintain homeostasis
hypoxia-inducible factor (HIF) are heterodimeric transcription factors whose activity is regulated by molecular oxygen through the prolyl hydroxylase domain (PHD) enzymes
We isolated uS12 as a binding partner of OGFOD1 and confirmed that the interaction is conserved between Ofd1 and Rps23 in fission yeast (Figure 1A; Figure 1—figure supplement 1A)
Summary
Eukaryotic cells require oxygen and must adapt to changes in its supply to maintain homeostasis. HIFs are heterodimeric transcription factors whose activity is regulated by molecular oxygen through the prolyl hydroxylase domain (PHD) enzymes. These enzymes hydroxylate HIF proline residues to promote HIF degradation using 2-oxoglutarate (2OG) and oxygen as co-substrates and Fe(II) as a co-factor (Kaelin and Ratcliffe, 2008). The PHDs belong to a large family of non-heme, 2OG/Fe(II)-dependent oxygenases whose members catalyze a broad set of reactions in eukaryotes and bacteria (Hausinger, 2015). Despite this diversity, the active site of most 2OG oxygenases is located in a highly-conserved double-stranded betahelix fold with the Fe(II) coordinated by a catalytic triad Since oxygen is a co-substrate for this enzyme family, they can act as sensors of cellular oxygen supply, making them ideal regulators for pathways requiring oxygen (Ratcliffe, 2013)
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