AbstractUnfolded protein response (UPR) plays an important role in the pathogenesis of many liver diseases. BMI1 has a liver protection effect, but whether it participates in the regulation of hepatocyte death through UPR is not well defined. Herein, the endoplasmic reticulum stress model was established by inducing hepatocyte line (MIHA) with tunicamycin (TM, 5 µg/ml). Cell counting kit-8 assay and flow cytometry were used to evaluate the viability and apoptosis of hepatocytes. The expression levels of BMI1, KAT2B, and proteins related to UPR (p-eIF2α, eIF2α, ATF4, and ATF6), NF-κB (p65 and p-p65), apoptosis (cleaved caspase-3, bcl-2, and bax) and necroptosis (p-MLKL and MLKL) were determined by Western blot. The relationship between KAT2B and BMI1 was determined by co-immunoprecipitation and ubiquitination assay. The results showed that TM not only promoted UPR, apoptosis, and necroptosis in hepatocytes but also upregulated the expression levels of BMI1 and KAT2B and activated NF-κB pathway. BAY-117082 reversed the effects of TM on viability, apoptosis, NF-κB pathway, and BMI1 but strengthened the effects of TM on KAT2B/MLKL-mediated necroptosis. BMI1 promoted the ubiquitination of KAT2B, and BMI1 overexpression reversed the effects of TM on viability, apoptosis, and KAT2B/MLKL-mediated necroptosis. In summary, overexpression of BMI1 promotes the ubiquitination of KAT2B to block the MLKL-mediated necroptosis of hepatocytes.