Synergistic interaction of triterpenoid saponins and plant protein toxins

Certain triterpenoid saponins from Gypsophila and Saponaria species augment the cytotoxicity of the type-I ribosome-inactivating protein (type-I RIPs) saporin from the seeds of Saponaria officinalis L. in a synergistic way. Type-I RIPs (25–30 kDa) are widely produced by Caryophyllaceae plants, which also synthesize triterpenoid saponins. The cytotoxic synergism between type I RIPs and saponins represents an evolutionarily conserved self-defense mechanism. To investigate the specificity of triterpenoid saponins in their ability to enhance the cytotoxicity of type-I RIPs we have isolated and characterized a triterpenoid saponin (SA1641) from Gypsophila paniculata L. SA1641 was combined with different type-I RIPs such as gelonin from Gelonium multiflorum A. Juss., agrostin from Agrostemma githago L., RIPQ3 from Stellaria media (L.) Vill. and saporin from Saponaria officinalis L. and cytotoxicity was determined in ECV-304 cells. Toxicity of saporin was drastically augmented by SA1641. Flow cytometric evaluations showed that this effect is not based on a SA1641-mediated induction of saporin endocytosis or a modulation of the plasma membrane permeability for saporin. We further confirmed that SA1641 does modulate the intracellular distribution of internalized saporin. Confocal microscopy indicated a SA1641-mediated release of saporin molecules out of cellular organelles (late endosome) and surface plasmon resonance measurements indicated an intracellular interaction of SA1641 and saporin as a prerequisite for the synergistic cytotoxicity between triterpenoid saponins and type-I RIPs.