| It is generally believed that ribosome-inactivating proteins (RIPs) are transported to their intracellular targets to express their toxicity. However, studies on the uptake, intracellular transport and apoptotic mechanism of type I RIPS and the cell-binding B chain of type II RIPS are rare. This study is to investigate some problems in these aspects of RIP toxicity.; We studied trichosanthin (TCS) and ricin A chain (RTA), which are type I RIPS, ricinus communis agglutinin (RCA), which is a type II RIP, and ricin B chain (RTB), which is the cell-binding chain of ricin and RCA.; TCS caused a decrease in cell number at the minimum effective dose of 800 nM at 24 hr post administration. The main type of cell death was apoptosis, which peaked at 12 hr.; RTA caused cell loss at the minimal dose of 50 nM at 24 hr. The main type of cell death was apoptosis, which peaked at 12 hr. RTA was internalised via a clathrin-dependent RME. Like the TCS transport, RTA was not found in the Golgi apparatus. The apoptosis proceeded via the extrinsic pathway that involved the activation of caspase-8 and caspase-3. However, on live rabbits, RTA caused necrotic skin damage.; RTB caused cell loss at the minimal dose of 100 nM at 24 hr. The main type of cell death was initially necrosis, but later became apoptosis, which peaked at 12 hr.; RCA caused a cell loss at the minimal dose of 50 nM at 24 hr. The main type of cell death was apoptosis, which peaked at 12 hr. The apoptosis proceeded through an extrinsic pathway that involved the activation of caspase-8, but not caspase-9.; These results show that (1) the cell-binding B chain is not a precondition for RIP toxicity, because TCS and RTA are also toxic to cells; (2) RTB itself is toxic; (3) without the binding of the B chain to cell surface, the entry and intracellular transport of type I RIPS differ from those of the type II; and (4) both RIPs and single B chain can induce apoptosis. Additionally, the results from live rabbits and cultured cells show that in vivo and in vitro toxicity may involve different cell death mechanisms. RTB-treated NIH 3T3 cells may serve as a model for the switch of cell death from necrosis to apoptosis. (Abstract shortened by UMI.)... |
