Functional evaluation of monomeric glutathione peroxidase 8 (Gpx8) as a mammalian redox sensor in the ER

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Thiol-disulfide mediated enzyme catalysis is not only essential for proper protein folding in the endoplasmic reticulum (ER), but is emerging to be involved in additional ER-related processes, such as ER stress sensing. ER stress sensing is a critical cellular event that controls unfolded protein response and associated cell death. Among the ER-resident thiol-dependent systems, peroxiredoxin-4 and two monomeric members of the glutathione peroxidase family of proteins, i.e. glutathione peroxidase 7 (Gpx7) and glutathione peroxidase 8 (Gpx8), are being considered to contribute to ER stress sensing via thiol-disulfide mediated exchange reactions. Mechanistically, Gpx7 increases the chaperone activity of GRP78/BiP (HSPA5, heat shock protein family A) by distinctive thiol-disulfide exchange reactions, and consequently Gpx7 knockout mice and cells accumulate misfolded proteins and show increased ER and oxidative stress. Nevertheless, Gpx7 shows very low and limited expression; therefore it is unlikely that it confers a more generalized role in sensing proper protein folding. By stark contrast, we found its related family member Gpx8 to be ubiquitously expressed in most tissues and cells with highest expression levels in kidney, heart, testis, adrenal gland and islet of pancreas. Additionally, we discovered that Gpx8 is dynamically regulated in response to ER and oxidative stress, although the underlying molecular mechanisms remain to be further explored. Moreover, we found that forced expression of Gpx8 renders cells resistant to fatty acid induced cell death indicating a link between fatty acid metabolism, Gpx8 and ER stress. To shed light into this barely studied member of the thiol-dependent redox network in the ER, we propose (i) to identify and functionally validate novel thiol-disulfide mediated redox partners of Gpx8, (ii) to interrogate the cellular mechanisms of Gpx8 loss in ER stress and associated cell death as well as the impact of the dynamic regulation of Gpx8 in response to ER stress, and (iii) as ultimate proof-of-concept to validate some of the key findings in novel mouse models proficient or deficient in Gpx8, along with a stress model of high fat diet induced metabolic syndrome and Type 2 diabetes mellitus. Hence, our studies aim at unravelling yet-unrecognized thiol-mediated ER stress sensing mechanisms, which will ultimately lead to new concepts in pathological conditions with an ER stress signature.