ATF2-mediated Transcriptional Activation of USP4 Stabilizes KMT2A Protein and Promotes Trophoblast Dysfunction

Preeclampsia (PE) is a severe and prevalent complication of pregnancy, characterized by insufficient trophoblast invasion, placental hypoxia, and systemic inflammation, which collectively represent the central pathological mechanisms underlying the disease. The molecular mechanism involving activating transcription factor 2 (ATF2), ubiquitin-specific peptidase 4 (USP4), and lysine methyltransferase 2 A (KMT2A) in hypoxia/reoxygenation (H/R)-treated trophoblast cells (HTR-8/SVneo) is explored, aiming to provide a novel theoretical basis for therapeutic strategies against PE. An in vitro H/R model was established using HTR-8/SVneo trophoblast cells to simulate PE pathophysiology. The expression levels of mRNA and protein were evaluated using RT-qPCR and western blotting, respectively. The proliferation, apoptosis, and invasion of cells were assessed using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), 5-ethynyl-2'-deoxyuridine (EdU), flow cytometry, and transwell assays. Ferroptosis-related markers were measured using commercial kits. The interaction between USP4 and KMT2A was determined by co-immunoprecipitation (Co-IP) and ubiquitination assays. Dual-luciferase reporter and chromatin immunoprecipitation (ChIP) assays were employed to investigate the binding of ATF2 to the USP4 promoter. KMT2A knockdown enhanced hypoxia/reoxygenation (H/R)-induced proliferation and invasiveness and suppressed apoptosis and ferroptosis in HTR-8/SVneo cells. The deubiquitinase USP4 stabilized KMT2A protein expression through deubiquitination. Overexpression of KMT2A reversed the impacts of USP4 silencing on H/R-mediated cellular injury and ferroptosis. Mechanistically, ATF2 transcriptionally activated USP4 expression. Furthermore, USP4 upregulation rescued the influences of ATF2 depletion on H/R-induced HTR-8/SVneo cell injury and ferroptosis. ATF2 activated USP4 transcription, which subsequently deubiquitinated and stabilized KMT2A protein expression, thereby influencing the progression of PE.