Low expression of cell-surface thromboxane A2 receptor β-isoform through the negative regulation of its membrane traffic by proteasomes

Human thromboxane A2 receptor (TP) consists of two alternatively spliced isoforms, TPα and TPβ, which differ in their cytoplasmic tails. To examine the functional difference between TPα and TPβ, we searched proteins bound to C termini of TP isoforms by a yeast two-hybrid system, and found that proteasome subunit α7 and proteasome activator PA28γ interacted potently with the C terminus of TPβ. The binding of TPβ with α7 and PA28γ was confirmed by co-immunoprecipitation and pull-down assays. MG-132 and lactacystin, proteasome inhibitors, increased cell-surface expression of TPβ, but not TPα. Scatchard analysis of [3H]SQ29548 binding revealed that the Bmax was higher in transiently TPα-expressing cells than TPα-expressing cells. In addition, TP-mediated phosphoinositide hydrolysis was clearly observed in TPα-, but not TPβ-expressing cells. These results suggest that TPβ binds to α7 and PA28γ, and the cell-surface expression of TPβ is lower than that of TPα through the negative regulation of its membrane traffic by proteasomes.