B chain is a functional subunit of β-bungarotoxin for inducing apoptotic death of human neuroblastoma SK-N-SH cells

Abstract

β-Bungarotoxin ( β-Bgt), a presynaptic phospholipase A 2 (PLA 2) neurotoxin isolated from the venom of Bungarus multicinctus, consists of A chain and B chain. The goal of the present study is to explore the functional contribution of the two subunits to the toxicity of β-Bgt. β-Bgt was found to induce apoptotic death of SK-N-SH cells via elevating intracellular Ca 2+ and intracellular ROS production. Moreover, an activation of p38 MAPK was associated with the cytotoxicity of β-Bgt. SB202190 (p38 MAPK inhibitor), N-acetylcysteine (antioxidant reagent), 1,2-bis(2-aminophenoxy)ethane- N, N, N, N-tetraacetic acid (BAPTA) (Ca 2+ chelator) and the inhibitors of Ca 2+ release from intracellular depots (ruthenium red and 2-aminoethoxydiphenyl borate) effectively attenuated the cytotoxicity of β-Bgt. In sharp contrast to the inability of A chain, B chain was able to induce cytotoxic effects on SK-N-SH cells as β-Bgt did. Abolishment of PLA 2 activity did not significantly alter the cytotoxic activity of β-Bgt. MK801 (an NMDA receptor antagonist), antibodies against NMDA receptor and 4-aminopyridine (a potassium channel blocker) markedly reduced the cytotoxic effects of β-Bgt, B chain and catalytically inactivated β-Bgt. Moreover, antibodies against NMDA receptor blocked the binding of rhodamine-labeled β-Bgt to SK-N-SH cells. Taken together, our data indicate that B chain is a functional subunit responsible for the cytotoxicity of β-Bgt, and suggest that the cytotoxicity of β-Bgt is mediated by NMDA receptor and potassium conductance.