Abstract
How basic polypeptide endocytosis got into living cells was a controversial and complex issue but with significant biological function. Cardiotoxins (CTXs), a major component of snake venom from Taiwan cobra Naja atra, are structurally homologues beta-sheet basic polypeptides that cause systolic cardiac arrest of the bitten victim with unknown internalization mechanism. (1) Previous study showed that CTX A2, A3 and A4 would get into cell and further exhibited toxicities; moreover, comparing amino acid between CTX A2 and A4, there are one amino acid difference. CTX A2 and A4 were specific to low or high sulfate heparan sulfate proteoglycans (HSPG), respectively. It is important to know what the endocytosis route of CTX A2, A3 and A4 is. According to these results, CTX A2 and A4 internalized via macropinocytosis and caveolae-mediated endocytosis at the same level but not for clathrin-mediated endocytosis. Therefore, one amino acid difference on protein caused different cell internalization responses in cells. (2) In the previous study, CTX A3 will cause pore-formation. Several researches show the correlation between membrane repair and internalization. We found that CTX A3 formed none-specific pore on cell membrane based on the result. In the presence of calcium, calcium influx from CTX A3-inducing pore triggered membrane repair process. However, in the absence of calcium, CTX A3 got into cell and targeted to mitochondria more obvious compared with in the presence of calcium environment. Besides calcium effect, cholesterol could also regulate CTX A3 internalization and localization. According to these data, basic polypeptide probably with different sensitive to HSPG and cholesterol caused different cell response. This indicated that protein internalization was regulated by multiple mechanisms and with high sensitive to HSPG and cholesterol of cell membrane.