Soapwort saponins trigger clathrin-mediated endocytosis of saporin, a type I ribosome-inactivating protein

Saporin, a type I ribosome-inactivating protein (RIP), removes adenine residues from the 28S ribosomal RNA as part of a process that leads to inhibition of protein synthesis. However, as shown in this study, neither saporin nor his-tagged saporin (both 0.6-6 pM) exert toxicity on several human cell lines including H-2171, SK-N-SH, HEP-G2, MOLT-3, THP-1, HL-60 and ECV-304. Saporin and his-tagged saporin became highly cytotoxic when they were used in a combined treatment with Soapwort saponins (SA). When combined with SA (2-4 μg/ml) saporin became as cytotoxic as the highly toxic type II RIP rViscumin reflected by an IC50 of 42.5 × 10−12 M for saporin and 21.5 × 10−12 M for rViscumin. We demonstrated that saporin was internalized via clathrin-mediated endocytosis, followed by the release into the endosomal transport system. Our results indicate that SA triggers this endocytic event rendering the otherwise cell membrane impermeable type I RIP saporin a potent cytotoxin. This effect was not cell line-specific suggesting that saporin exploits a common SA-dependent mechanism to enter cells.