KRT20 Protects against Ferroptosis of Renal Tubular Cells by Inhibiting Extracellular Secretion of PRDX2 in AKI

Background: Acute kidney injury (AKI) exhibits high incidence and morbidity. Ferroptosis of renal tubular cells is a key event driving tubular injury in AKI, but the mechanism of which is unclear. Cytokeratin 20 (KRT20), a biomarker of tubular injury, is an intermediate filament protein whose role in renal tubular injury remain elusive. Intracellular peroxiredoxin2(PRDX2) is closely related to ferroptosis, but the relationship between KRT20 and PRDX2 has not been reported in any studies. Methods: Bulk RNA-seq and snRNA-seq of kidneys from the AKI mouse model in public databases, were used to analyze transcript levels of KRT20 in AKI, which was validated in two AKI mouse models (cisplatin-induced AKI and ischemia-reperfusion(IR) AKI) and clinical kidney biopsy specimens. Renal proximal tubule-specific Krt20 knockout mice in vivo, knockdown and overexpression of KRT20 in vitro, were used to construct two AKI models to assess the role of KRT20 in AKI. Luciferase reporter assays, CHIP-PCR and IP-MS were used to explore the transcription factors and the downstream candidates target of KRT20, respectively. CO-IP and truncating mutation were used to explore the binding domain between KRT20 and candidate targets. Whether urinary PRDX2 could be used as a promising predictor for AKI was investigated in a patient cohort. Results: KRT20 was dramatically increased in renal tubules upon injury in the AKI mouse model and AKI patients, which might be regulated by the transcription factor Fosb. The renal tubular specific knockout of Krt20 aggravated cisplatin- and IR-induced AKI in mice. The intervention of KRT20 affected ferroptosis of tubular cells in vitro. Mechanistically, KRT20 bound with PRDX2, whose secretion was dependent on ALG-2-interacting protein X(Alix), to retain its intracellular location to prevent renal tubular cell ferroptosis and attenuate AKI. KRT20 and Alix competitively bind to the NTD domain of PRDX2, thus affecting the secretion of PRDX2. Urinary concentrations of PRDX2 significantly increased in patients who underwent partial nephrectomy, which positively correlated with the renal tubular injury biomarker, such as KIM-1 and NGAL. Conclusion: This study discovered a Fosb-KRT20-PRDX2 axis in the progression of AKI, which warrants future exploration as a therapeutic targets. In addition, PRDX2 may be a noninvasive biomarker for renal tubular injury. Funding: Government Support – Non-U.S.