Berberrubine alleviates PCS-induced tubular injury via HNRNPF/NR4A1/ER stress axis

The key uremic toxin p-cresyl sulfate (PCS) has been shown to induce renal tubule injury-mediated chronic kidney disease via endoplasmic reticulum stress (ERS). Berberine offers renoprotective benefits and attenuates ERS-driven apoptosis. Owing to the low oral bioavailability of berberine, its primary metabolite, berberrubine (BRB), here considered a direct bioactive-agent responsible for its effects. However, the protective effects of BRB against nephritic damage and its regulatory function involving ERS or underlying molecular targets remain poorly understood. In this work, biochemical, cellular assays and animal studies revealed that BRB alleviates PCS-induced renal dysfunction, pathological damage, ERS and apoptosis. Additionally, transcriptomics, biochemical and cellular assays revealed that BRB effectively inhibited the PCS-mediated phosphorylation and nuclear translocation of NR4A1, which in turn suppressed the GRP78/IRE1α/XBP1s/CHOP signaling pathway. Mechanistically, DARTS, CETSA, SPR and mutant plasmids identified HNRNPF as the functional target of BRB, with key binding sites TYR306 and ASN313, further revealing its role as a critical regulator of the NR4A1/GRP78/IRE1α/XBP1s/CHOP signaling pathway in HNRNPF-knockdown/overexpressing HK-2 cells and AAV9-saCAS9-sgRNA knockdown C57BL/6 mice. Our findings reveal a previously unrecognized HNRNPF/NR4A1/ERS pathway in which BRB targets HNRNPF as a newly identified small-molecule ligand, and these results offer valuable insights into the potential targeting of HNRNPF for the treatment of tubule lesions.