Abstract
Cobra cardiotoxins (CTXs) are structurally related snake venom beta-sheet polypeptides that cause systolic cardiac arrest and severe tissue necrosis of the bitten victim with unknown mechanism. Here, we show that sulfatide (SGC) lipid domain is involved in CTX A3 action in H9C2 cell not only to promote CTX pore formation in plasma membrane but also to facilitate CTX internalization for targeting mitochondria. SGC can interact with CTX A3 to form a non-specific ion conduction pore on cell membrane via a sulfatide-induced insertion of CTX into cell membrane. The CTX-induced lipid conformational change at the ceramide interfacial region is a key step for the action of CTX A3 to induce oligomerization and to cause its cytotoxicity in an SGC-dependent manner. CTX A3 internalization is facilitated by sulfatide in plasma membrane by a cholesterol-sensitive, temperature-dependent, ATP-independent, caveolae-independent, and clathrin-independent, endocytic pathway. Based on exogenous addition of heparin with various chain-lengths, we propose that CTX A3 dimerization is an important intermediate for internalization. CTX A3 targets mitochondria after its internalization in ~5 minutes and disrupts the mitochondrial network in the presence or absence of CTX-induced calcium increase. By action on the plasma and mitochondrial membranes, CTX-induced a necrotic cell death of H9C2 myoblasts.