The protein toxin ricin is found in the plant Ricinus communis, and is a useful tool in studies of intracellular traffic. It consists of an enzymatically active A chain and a B chain with lectin properties, joined together by a disulfide bond. Ricin is transported from the plasma membrane, via the Golgi apparatus, and to the endoplasmic reticulum. The A chain is then translocated to the cytosol, where it inactivates ribosomes and thus inhibits protein synthesis. Using a modified form of ricin containing a tyrosine sulfation site (ricin sulf-1) makes it possible to measure the amount of ricin transported to the Golgi apparatus, as sulfation is a Golgi modification.
In this study, we have looked into the role played by the phosphatidyl inositol 3-kinase hVps34 in endosome to Golgi retrograde transport of ricin. The effect has been studied by transiently transfecting HEK cells with wildtype hVps34, dominant negative mutants of hVps34 and vector based siRNA directed against hVps34. We have also made use of the PI 3-kinase inhibitors wortmannin and LY294002. The results from biochemical approaches together with confocal microscopy indicate that hVps34 is indeed involved in endosome to Golgi transport of ricin.