Abstract
We demonstrated recently that in Chinese hamster ovary cells stably expressing human recombinant endothelinA receptors (CHO-ETAR), endothelin-1 (ET-1) activates two types of Ca2+-permeable nonselective cation channels (designated NSCC-1 and NSCC-2) and a store-operated Ca2+ channel (SOCC), which can be distinguished by Ca2+ channel blockers such as 1-{β-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenylethyl}-1H-imidazole hydrochloride (SK&F 96365) and (R,S)-(3,4-dihydro-6,7-dimethoxy-isochinolin-1-yl)-2-phenyl-N,N-di[2-(2,3,4-trimethoxyphenyl)ethyl]acetamid mesylate (LOE 908). We also reported that CHO-ETAR couples with G12 in addition to Gq and Gs. The purpose of the present study was to identify the G proteins involved in the activation of these Ca2+ channels by ET-1, using mutated ETARs with coupling to either Gqor Gs/G12 (designated ETARΔ385 and SerETAR, respectively) and a dominant-negative mutant of G12 (G12G228A). ETARΔ385 is truncated immediately downstream of Cys385 in the C terminus as palmitoylation sites, whereas SerETAR is unpalmitoylated because of substitution of all the cysteine residues to serine (Cys383Cys385–388 → Ser383Ser385–388). In CHO-ETARΔ385, stimulation with ET-1 activated only SOCC. In CHO-SerETAR or CHO-ETAR pretreated withU73122, an inhibitor of phospholipase C (PLC), ET-1 activated only NSCC-1. Dibutyryl cAMP alone did not activate any Ca2+channels in the resting and ET-1–stimulated CHO-SerETAR. Microinjection of G12G228A abolished the activation of NSCC-1 and NSCC-2 in CHO-ETAR and that of NSCC-1 in CHO-SerETAR. These results indicate that ETAR activates three types of Ca2+ channels via different G protein-related pathways. NSCC-1 is activated via a G12-dependent pathway, NSCC-2 via Gq/PLC- and G12-dependent pathways, and SOCC via a Gq/PLC-dependent pathway.
- The American Society for Pharmacology and Experimental Therapeutics
MolPharm articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|