Nephrol Dial Transplant. 2026 Sep 2:gfag202. doi: 10.1093/ndt/gfag202. Online ahead of print.
ABSTRACT
BACKGROUND: Chronic kidney disease (CKD) is characterized by proximal tubule (PT) stress, oxidative injury, and metabolic dysfunction. Human kidney single-nucleus RNA-sequencing (snRNA-seq) identified enrichment of ubiquitin-dependent protein catabolic processes in injured PT cells, suggesting activation of the ubiquitin-proteasome system during tubular stress. Because Cullin 3 (CUL3), scaffold of ubiquitin ligases, regulates oxidative stress signaling through the KEAP1-NRF2 axis, we investigated its role in PT injury and stress adaptation.
METHODS: Human CKD snRNA-seq data and kidney immunostaining were used to define PT cell states, CUL3-associated pathways, and CUL3 localization. Injury-associated CUL3 regulation was examined in wildtype mice after ischemia-reperfusion injury (IRI). Inducible PT-specific knockout mice (Slc34a1-CreER; Cul3flox/flox) were analyzed at baseline and after injury by histology, immunostaining, proteomics, and injury assessment. In addition, proteomic analysis of a whole-tubule epithelial knockout model (Pax8-rtTA/LC1; Cul3flox/flox) was performed. In immortalized human PT cells, CUL3 was suppressed or activated using CRISPR interference and CRISPR activation, followed by bulk RNA sequencing.
RESULTS: CUL3 transcript and protein expression was enriched in stressed PT states in human CKD. In mice, CUL3 protein abundance increased after injury, supporting injury-associated induction in vivo. PT-specific CUL3 deletion increased antioxidant NQO1 expression without causing overt baseline injury. Proteomic analysis of isolated CUL3-deficient PT cells revealed induction of antioxidant, detoxification, proteostasis, and lipid metabolic programs, together with suppression of mitochondrial oxidative metabolism. Similar changes were observed in whole-tubule Cul3 knockout model. In gene-edited human PT cells, CUL3 suppression recapitulated stress-associated and metabolic remodeling programs, whereas CUL3 activation induced reciprocal transcriptional changes. Despite induction of antioxidant pathways, PT-specific CUL3 deletion did not alter disease severity after IRI or aristolochic acid nephropathy.
CONCLUSION: CUL3 is an injury-induced regulator of PT metabolic and stress-associated states and modulates antioxidant defense and mitochondrial metabolism in PT cells.
PMID:42684049 | DOI:10.1093/ndt/gfag202