Regulation and intracellular trafficking pathways of the endothelin receptors

The effects of endothelin (ET) are mediated via the G protein-coupled recep tors ETA and ETB. However, the mechanisms of ET receptor desensitization, i nternalization, and intracellular trafficking are poorly understood. The ai m of the present study was to investigate the molecular mechanisms of ET re ceptor regulation and to characterize the intracellular pathways of ET-stim ulated ETA and ETB receptors, By analysis of ETA and ETB receptor internali zation in transfected Chinese hamster ovary cells in the presence of overex pressed beta ARK, beta-arrestin-1, beta-arrestin-2, or dynamin as well as d ominant negative mutants of these regulators, we have demonstrated that bot h ET receptor subtypes follow an arrestin- and dynamin/clathrin-dependent m echanism of internalization. Fluorescence microscopy of Chinese hamster ova ry and COS cells expressing green fluorescent protein (GFP)-tagged ET recep tors revealed that the ETA and ETB subtypes were targeted to different intr acellular routes after ET stimulation. While ETB-GFP followed a recycling p athway and colocalized with transferrin in the pericentriolar recycling com partment, ETB-GFP was targeted to lysosomes after ET-induced internalizatio n. Both receptor subtypes colocalized with Rab5 in classical early endosome s, indicating that this compartment is a common early intermediate for the two ET receptors during intracellular transport. The distinct intracellular routes of ET-stimulated ETA and ETB receptors may explain the persistent s ignal response through the ETA receptor and the transient response through the ETB receptor. Furthermore, lysosomal targeting of the ETB receptor coul d serve as a biochemical mechanism for clearance of plasma endothelin via t his subtype.