Inhibition of Biosynthesis of Human Endothelin B Receptor by the Cyclodepsipeptide Cotransin

The specific inhibition of the biosynthesis of target proteins is a relatively novel strategy in pharmacology and is based mainly on antisense approaches (e.g. antisense oligonucleotides or RNA interference). Recently, a novel class of substances was described acting at a later step of protein biosynthesis. The cyclic heptadepsipeptides CAM741 and cotransin were shown to inhibit selectively the biosynthesis of a small subset of secretory proteins by preventing stable insertion of the nascent chains into the Sec61 translocon complex at the endoplasmic reticulum membrane (Besemer, J., Harant, H., Wang, S., Oberhauser, B., Marquardt, K., Foster, C. A., Schreiner, E. P., de Vries, J. E., Dascher-Nadel, C., and Lindley, I. J. (2005) Nature 436, 290-293; Garrison, J. L., Kunkel, E. J., Hegde, R. S., and Taunton, J. (2005) Nature 436, 285-289). These peptides act in a signal sequence-discriminatory manner, which explains their selectivity. Here, we have analyzed the cotransin sensitivity of various G protein-coupled receptors in transfected HEK 293 cells. We show that the biosynthesis of the human endothelin B receptor (ETBR) is highly sensitive to cotransin, in contrast to that of the other G protein-coupled receptors analyzed. Using a novel biosynthesis assay based on fusions with the photoconvertible Kaede protein, we show that the IC50 value of cotransin action on ETBR biosynthesis is 5.4 mu M and that ETBR signaling could be completely blocked by treating cells with 30 mu M cotransin. Taken together, our data add an integral membrane protein, namely the ETBR, to the small group of cotransin-sensitive proteins.