Evolutionary Insights into Oxygen Sensing

In animals, the hypoxia‐inducible factor (HIF) system is important both in sensing oxygen levels and in coordinating cellular responses to hypoxia. Factor inhibiting HIF (FIH) and the HIF prolyl‐hydroxylases (PHD 1–3) are Fe(II) and 2‐oxoglutarate (2OG) dependent oxygenases that act as oxygen sensors using different, but related mechanisms. They catalyse the hydroxylation of asparagine‐ or proline‐residues of HIF‐α, in order to reduce the activity or levels of HIF‐α, respectively. Reduced HIF‐α hydroxylase activity, in turn, upregulates the expression of genes that work to counteract the effects of hypoxia. The evolutionary origins of the HIF sensing system before animals are unclear. However, recent work has suggested that prokaryotic prolyl‐hydroxylases involved in the regulation of translation may have predated the HIF prolyl‐hydroxylases. In Dictyostelium discoideum , a protistan slime mould, prolyl 4‐hydroxylase‐1 (DdP4H1) has been reported to have a role in oxygen sensing; interestingly, no HIF exists in Dictyostelium . Instead, the E3‐ubiquitin ligase adaptor S‐phase kinase‐associated protein 1 (Skp1) is the substrate of DdP4H1. DdSkp1 is subject to hydroxylation on Pro 143 to produce hydroxylated Skp1 (HO‐Skp1). HO‐Skp1 is then attached to a Galα1–6Galα1–3Fucα1–2Galβ1–3GlcNAc pentasaccharide, by glycosyl transferases that regulate the expression of various genes. Toxoplasma gondii , a protozoan parasite, possesses a similar oxygen sensing system as Dictyostelium . It is proposed that TgSkp1 hydroxylation is catalysed by TgPhyA in a manner dependent on the oxygen level. Skp1 also exists in humans. However, the equivalent residue to the proline in Dictyostelium or Toxoplasma , is a Glutamate in human Skp1 which is not hydroxylated. This research is concerned with biochemical studies on the evolution of the HIF and related oxygen sensing systems, focusing on the roles and properties of the prolyl‐hydroxylases and their reactions with oxygen. Such studies may help to identify new roles for the PHDs as yet unidentified in human cells. Biochemical and biophysical analysis on P4H‐Skp1 or PhyA‐Skp1 has been carried out to investigate their possible roles as oxygen sensors in Dictyostelium and Toxoplasma . It is aimed to compare their kinetic properties especially in terms of oxygen binding reactions with that of the human PHDs. A second aim is to track the evolution of the oxygen sensing mechanism in eukaryotes to identify the origins of hypoxia sensing in animals. Support or Funding Information Department of Chemistry, University of Oxford, Chemistry Research Laboratory, 12 Mansfield Road, Oxford, OX1 3TA, UKP4H catalysed hydroxylation. P4H rely on oxygen and Fe(II) to couple substrate two electron oxidation to the decarboxylation of 2OG to succinate and carbon dioxide. PPHD and human HIF (PHDs), catalyse trans ‐4‐prolyl‐hydroxylation.