The C-terminal region regulates the constitutive signaling activity of GPR119: G protein recruitment

G protein-coupled receptor 119 (GPR119), a glucose-dependent insulinotropic receptor, modulates glucose homeostasis. The complexity and diversity of GPR119 signaling, closely associated with its C-terminal domain, may hinder the development of selective drugs and contribute to off-target effects. This study systematically examined the function of the C-terminal domain using amino acid deletions and site-directed mutagenesis, assessing cyclic adenosine monophosphate (cAMP) accumulation, the recruitment of G_s protein, β-arrestin 1 (βarr1), and β-arrestin 2 (βarr2). We initially conducted a broad screening through truncations of 9, 18, 27, and 36 amino acids, followed by targeted single-amino-acid deletion analyses. These results revealed that the 15-amino-acid truncation was essential for G_s protein recruitment and cAMP accumulation. Protein purification results revealed that truncation of the C-terminal domain did not affect GPR119-G_s-G_βγ complex formation. We further demonstrated that the C-terminal proximal phosphorylation motif was essential for the recruitment of G_s protein, βarr1, and βarr2. Notably, phosphorylation motifs in the C-terminal domains of Trace Amine-Associated Receptor 1 (TAAR1) and β-2 Adrenergic Receptor (β₂AR) displayed distinct differences in the G_s protein-mediated cAMP accumulation. These findings improve our understanding of GPCR signaling's diversity and complexity, offering new insights for the development of GPR119-targeted therapeutics.