Oxygen-dependent formation of stable protein oligomers (oxomers) - a novel signal transduction in oxygen (patho-)physiology

• Applicant: Professor Dr. Carsten Scholz
• Subject Area: Anatomy and Physiology
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 528713131

Oxygen depletion (hypoxia) occurs under both physiological and pathophysiological conditions, including exercise, inflammation and fibrosis. Human cells are capable of sensing changes in local oxygen supply, allowing the cells to adapt to hypoxia. The major cellular oxygen sensors are prolyl-4-hydroxylase domain (PHD) proteins 1-3 and factor inhibiting HIF (FIH). PHDs and FIH hydroxylate hypoxia-inducible factor (HIF) α subunits, which reduces HIF-α half-life (PHDs) or modulates HIF transactivation activity (FIH), respectively. The PHDs and FIH are likely also relevant for oxygen (patho-)physiology independent of HIF, but the underlying mechanism(s) are largely unclear. Six separate pharmacologic HIF hydroxylase inhibitors (HIs) have recently been approved for the treatment of patients with renal anemia. These drugs will likely also be used for tissue protection in hypoxia-associated diseases, such as inflammation and fibrosis. However, the underlying mechanisms of the tissue protective HI effect are mainly unknown, hindering their translation into the clinics for such treatments. Therefore, it is of utmost importance to improve our understanding of (patho-)physiological functions of the PHDs and FIH, especially outside the HIF pathway. We recently found that FIH forms an “oxomer” – defined as oxygen-dependent stable protein oligomer – with the deubiquitinase OTUB1 (FIH-OTUB1). Furthermore, we identified with a newly developed mass spectrometry (MS)-based approach 12 additional putative FIH-dependent oxomers with other proteins. FIH-OTUB1 oxomer formation regulates OTUB1 enzymatic activity, thus representing a novel mode of oxygen-dependent cellular signaling. The goal of this application is to characterize the occurrence, regulation and function of oxomers. Subproject A will analyze the mechanism of a previously unknown pro-inflammatory increase of FIH and OTUB1 protein levels (unpublished data) as well as its effect on the FIH-OTUB1 oxomer in renal cells and tissues. Subsequently, the impact of the FIH-OTUB1 oxomer on pro-inflammatory and pro-fibrotic signaling pathways will be assessed. Subproject B will focus on PHD-dependent oxomer formation and function. Utilizing the previously established MS-based analysis, our unpublished data shows that all three PHDs form oxomers with several different target proteins. In this subproject, we will verify the identified oxomers and assess the involved amino acid residues in the oxomer formation, the mechanism of formation and formation kinetics, half-life and oxygen sensitivity as well as the function of selected oxomers. The characterization and analysis of this novel type of oxygen-dependent signal transduction via oxomer formation may lead to a paradigm shift of the role of PHDs and FIH in oxygen (patho-)physiology and will improve our understanding of the mechanism of action of HIs.

• DFG Programme: Research Grants