The involvement of BNIP3 in cyanide-induced neurotoxicity

Lu Zhang, Purdue University

Abstract

In this study, an immortalized dopaminergic cell line was used to characterize the cell death signaling cascade activated by cyanide. Cyanide-treated cells exhibited a time- and concentration-dependent apoptosis that was caspase-independent. Cyanide induced a rapid surge of intracellular reactive oxygen species (ROS) generation, followed by p38 mitogen-activated protein kinase (MAPK) activation and nuclear accumulation of hypoxia-inducible factor-1α (HIF-1α). Cyanide activated the hypoxia response element (HRE) promoter, followed by increased BNIP3 gene transcription. BNIP3 upregulation was reduced by selective RNAi knockdown of HIF-1α. Overexpression of BNIP3 produced mitochondrial dysfunction, caspase-independent apoptosis, and sensitization of the cells to cyanide-induced toxicity. It was concluded that cyanide activated the HIF-1α-mediated pathway of BNIP3 induction through a redox-sensitive process. Increased BNIP3 expression then served as an initiator of mitochondria-mediated death. In mice, repeated exposure to a sublethal dose of cyanide resulted in an increase of BNIP3 expression in midbrain, which was accompanied by a loss of dopaminergic neurons. In cortex, cyanide induced upregulation of BNIP3, which may be involved in cortical cell apoptosis. ^ In the current study, Ca²⁺ mobilization between endoplasmic reticulum (ER) and mitochondria in BNIP3-mediated cell death process was determined. BNIP3 was localized in both ER and mitochondria to facilitate the release of Ca²⁺ from ER and subsequently increase uptake of Ca ²⁺ into mitochondria. Excessive accumulation of mitochondrial Ca ²⁺ facilitated the loss of ΔΨ_m, ultimately resulting in the execution of cell death. The alteration of Ca²⁺ in ER and mitochondria can be blocked by anti-apoptotic protein, Bcl-2. In addition, mitochondria-targeted BNIP3 initiates cell apoptosis in a Ca²⁺-indepenent mechanisms by inducing MPT and ΔΨ_m dissipation. This process cannot be blocked by Bcl-2 overexpression. Current results also confirm that Bax plays a critical role in regulating the mitochondria dysfunction produced by BNIP3. In response to cyanide, Bax was activated and co-localized with BNIP3 in ER and mitochondria. Forced overexpression of BNIP3 also resulted in the activation of Bax. Acting as a downstream effector for BNIP3, Bax is required for Ca²⁺ crosstalk between ER and mitochondria, thereby contributing to mitochondria-mediated cell apoptosis.^

Degree

Ph.D.

Advisors

Gary E. Isom, Purdue University.

Subject Area

Health Sciences, Pharmacology

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