GRINA alleviates hepatic ischemia‒reperfusion injury-induced apoptosis and ER-phagy by enhancing HRD1-mediated ATF6 ubiquitination

Abstract

Background & aims: Hepatic ischemia-reperfusion injury (HIRI) is a critical complication of liver surgery and transplantation that contributes significantly to severe organ failure. GRINA, a calcium-regulating endoplasmic reticulum (ER) protein, plays an essential role in controlling the unfolded protein response; however, its role in HIRI remains unclear. The aim of this study was to investigate the function of GRINA in HIRI and explore its potential as a therapeutic target.

Methods: Liver tissues from patients undergoing hepatectomy, alongside a mouse model of partial HIRI, were used to assess GRINA expression levels. Hepatocyte-specific Grina knockout and transgenic mouse models were generated to explore the effects of GRINA on HIRI. Key markers of inflammation, apoptosis, ER stress, and autophagy were evaluated via real-time PCR, Western blotting, immunohistochemistry, immunofluorescence, and ELISA. RNA sequencing, mass spectrometry, coimmunoprecipitation and ubiquitination assays were used to elucidate the underlying molecular mechanisms.

Results: GRINA expression was markedly reduced in hepatocytes from both patients and mice with HIRI, and its expression was inversely correlated with the severity of liver damage. Hepatocyte-specific Grina overexpression mitigated liver injury, the inflammatory response, and hepatocyte apoptosis following HIRI, whereas GRINA deficiency exacerbated these outcomes. Mechanistically, GRINA interacted directly with ATF6 and recruited HRD1 to form a multiprotein complex that catalyzed ATF6 polyubiquitination, thereby promoting its degradation. This process suppressed ER autophagy (ER-phagy), providing cellular protection following HIRI. The inhibition of ATF6 degradation attenuated the protective effects of GRINA in HIRI.

Conclusions: Our study highlights the critical role of the GRINA-HRD1-ATF6 complex in regulating ER stress and autophagy during HIRI. These findings provide new insights into therapeutic strategies to alleviate HIRI.

Impact and implications: Hepatic ischemia-reperfusion injury (HIRI) represents a multifaceted pathophysiological challenge commonly encountered during liver surgeries, yet its underlying molecular mechanisms remain inadequately understood. In this study, we revealed a significant negative correlation between GRINA levels and the severity of liver damage in patients with HIRI. Our findings demonstrate that GRINA alleviates endoplasmic reticulum stress by enhancing HRD1-mediated ubiquitination of ATF6, thereby maintaining calcium homeostasis and inhibiting ER-phagy. This study provides novel insights into the role of GRINA in protecting liver cells under HIRI, offering fresh perspectives for clinical prevention and management strategies for HIRI.

Keywords: ATF6; Endoplasmic reticulum stress; GRINA; Hepatic ischemia‒reperfusion injury; Ubiquitination; autophagy.