Human chorionic gonadotropin-ricin A chain hybrid protein: a hormone analog for the study of signal transduction.

We have prepared a disulfide cross-linked hybrid consisting of human chorionic gonadotropin and the toxic ricin A chain, utilizing N-succinimidyl-3-(2-pyridyldithio)-propionate. The purified hybrid exhibited cytotoxicity toward rat testis Leydig cells due to inhibition of protein synthesis within the cells, although it inhibited protein synthesis in a cell-free system of a rabbit reticulocyte lysate only after treatment of the hCG-A hybrid with dithiothreitol. On the other hand, pretreatment of the hCG-A hybrid with dithiothreitol completely abolished its cytotoxicity. The hCG-A hybrid stimulated the testosterone production by rat testis Leydig cells to a similar degree to hCG. The hCG-A hybrid could elicit full testosterone production in the presence of about 1/3-1/2 less accumulated cAMP compared with in the case of hCG stimulation. The steroidogenetic effect of the hCG-A hybrid was largely inhibited by anti-hCG antibody but not by anti-ricin A chain antibody. These results suggest that the hCG-A hybrid first binds to Leydig cells via an hCG receptor and then is internalized into the cells, and finally the inhibition of cellular protein synthesis occurs on the release of the ricin A chain from the hybrid. Furthermore, it was suggested that hCG may stimulate testosterone production via another second messenger system as well as via the cAMP system, since the hCG-A hybrid stimulated full testosterone production without maximal cAMP accumulation.