A study of Marchf6 as an E3 ubiquitin ligase for pro-opiomelanocortin (POMC) degradation

Abstract

The metabolic prohormone pro-opiomelanocortin (POMC) is typically translocated into the endoplasmic reticulum (ER) for entry into the secretory pathway. Patients harboring mutations within the signal peptide (SP) of POMC or its adjoining segment develop metabolic disorders. However, the existence, metabolic fate, and functional consequences of cytosol-retained POMC remain elusive. In the first part of this study, I found that Marchf6 as an ER-embedded E3 ligase targets cytosolic POMC in hypothalamic POMC neuronal cells. Notably, SP-uncleaved POMC is recognized by Marchf6, while, the proline 460 residue in the cytosolic 4th domain (C4) of Marchf6 plays a pivotal role in both POMC recognition and degradation via the ubiquitin-proteasome system (UPS). In the other part of this study, I unveiled that the upregulation of cytosol-retained POMC induces ER stress and ferroptosis, a form of cell death associated with iron- and lipid reactive oxygen species (ROS). Mechanistically, cytosol-retained POMC sequesters the chaperone Hspa5, thereby accelerating the degradation of glutathione peroxidase Gpx4, a key regulator of ferroptosis, through chaperone-mediated autophagy. Interestingly, the ER membrane-embedded Marchf6 E3 ubiquitin ligase facilitates the proteasomal destruction of cytosol-retained POMC, thereby preventing ER stress and ferroptosis. I also found that Marchf6 suppresses the PERK/eIF2α/Aft4-related ER stress response and the Nox2/Nox4 NADPH oxidase-mediated ROS production. In the last phase of this study, I generated that POMC neuron-specific Marchf6-deficient mice (Marchf6POMC) to clarify the physiological implications of Marchf6-mediated POMC degradation in animal model. Employing diverse metabolic analyses and immunohistochemistry approaches, remarkably, I observed a significant upregulation of ER stress markers and an elevation in lipid ROS levels, accompanied by concurrent reductions in Gpx4 levels in the hypothalamic arcuate nucleus (ARC) of Marchf6POMC mice. Consequently, Marchf6POMC exhibited increased body weight, hyperphagia, and attenuated energy expenditure. Collectively, I elucidated that POMC is a previously unrecognized substrate of Marchf6. Moreover, I provided compelling evidence supporting the role of Marchf6 in modulating ferroptosis through the degradation of cytosolic-retained POMC. Furthermore, I confirmed POMC neuron-specific Marchf6-deficient mice suffer from hyperphagia, weight gain, and decreased energy expenditure partly due to upregulated POMC-mediated ferroptotic damage and ER stress. Taken together, the comprehensive findings of this study firmly establish Marchf6 as a critical regulator of ER stress, ferroptosis, and metabolic homeostasis within the realm of POMC neurons.