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Bax inhibitor-1 (BI-1) is an evolutionarily conserved protein that protects cells against endoplasmic reticulum (ER) stress while also affecting the ER stress response. In this study, we examined BI-1-induced regulation of the ER stress response as well as the control of the protein over cell death under ER stress. In BI-1-overexpressing cells (BI-1 cells), proteasome activity was similar to that of control cells; however, the lysosomal fraction of BI-1 cells showed sensitivity to degradation of BSA. In addition, areas and polygonal lengths of lysosomes were greater in BI-1 cells than in control cells, as assessed by fluorescence and electron microscopy. In BI-1 cells, lysosomal pH was lower than in control cells and lysosomal vacuolar H+-ATPase(V-ATPase), a proton pump, was activated, suggesting high H+ uptake into lysosomes. Even when exposed to ER stress, BI-1 cells maintained high levels of lysosomal activities, including V-ATPase activity. Bafilomycin, a V-ATPase inhibitor, leads to the reversal of BI-1-induced regulation of ER stress response and cell death due to ER stress. In BI-1 knock-out mouse embryo fibroblasts, lysosomal activity and number per cell were relatively lower than in BI-1 wild-type cells. This study suggests that highly maintained lysosomal activity may be one of the mechanisms by which BI-1 exerts its regulatory effects on the ER stress response and cell death. [ABSTRACT FROM AUTHOR]

The POZ domain is a highly conserved protein-protein interaction motif found in many regulatory proteins. Nuclear factor-κB (NF-κB) plays a key role in the expression of a variety of genes in response to infection, inflammation, and stressful conditions. We found that the POZ domain of FBI-1 (factor that binds to the inducer of short transcripts of human immunodeficiency virus-1) interacted with the Rel homology domain of the p65 subunit of NF-κB in both in vivo and in vitro protein-protein interaction assays. FBI-1 enhanced NF-κB-mediated transcription of E-selectin genes in HeLa cells upon phorbol 12-myristate 13-acetate stimulation and overcame gene repression by IκBα or IκBβ. In contrast, the POZ domain of FBI-1, which is a dominant-negative form of FBI-1, repressed NF-κB-mediated transcription, and the repression was cooperative with IκBα or IκBβ. In contrast, the POZ domain tagged with a nuclear localization sequence polypeptide of FBI-1 enhanced NF-κB-responsive gene transcription, suggesting that the molecular interaction between the POZ domain and the Rel homology domain of p65 and the nuclear localization by the nuclear localization sequence are important in the transcription enhancement mediated by FBI-1. Confocal microscopy showed that FBI-1 increased NF-κB movement into the nucleus and increased the stability of NF-κB in the nucleus, which enhanced NF-κB-mediated transcription of the E-selectin gene. FBI-1 also interacted with IκBα and IκBβ. [ABSTRACT FROM AUTHOR]