Role of the differentially spliced carboxyl terminus in thromboxane A(2) receptor trafficking - Identifcation of a distinct motif for tonic internalization

The thromboxane A(2) receptor (TP) is a G protein-coupled receptor that is expressed as two alternatively spliced isoforms, alpha (343 residues) and b eta (407 residues) that share the first 328 residues. We have previously sh own that TP beta, but not TP alpha, undergoes agonist-induced internalizati on in a dynamin-, GRK-, and arrestin-dependent manner. In the present repor t, we demonstrate that TP beta, but not TP alpha, also undergoes tonic inte rnalization. Tonic internalization of TP beta was temperature- and dynamin- dependent and was inhibited by sucrose and NH4Cl treatment but unaffected b y wildtype or dominant-negative GRKs or arrestins. Truncation and site-dire cted mutagenesis revealed that a YX(3)phi moth (where X is any residue and phi is a bulky hydro phobic residue) found in the proximal portion of the c arboxyl-terminal tail of TP beta was critical for tonic internalization but had no role in agonist-induced internalization. Interestingly, introductio n of either a YX(2)phi or YX(3)phi motif in the carboxyl-terminal tail of T P alpha induced tonic internalization of this receptor. Additional analysis revealed that tonically internalized TP beta undergoes recycling back to t he cell surface suggesting that tonic internalization may play a role in ma intaining an intracellular pool of TP beta, Our data demonstrate the presen ce of distinct signals for tonic and agonist-induced internalization of TP beta and represent the first report of a YX(3)phi motif involved in tonic i nternalization of a cell surface receptor.