Characterization of the carboxyl-terminal domain of the rat glucose-dependent insulinotropic polypeptide (GIP) receptor - A role for serines 426 and 427 in regulating the rate of internalization

Glucose-dependent insulinotropic polypeptide (GIP) is a gastrointestinal ho rmone involved in the regulation of insulin secretion. In non-insulin-depen dent diabetes mellitus insulin responses to GIP are blunted, possibly due t o altered signal transduction or reduced receptor number. Site-directed mut agenesis was used to construct truncated GIP receptors to study the importa nce of the carboxyl-terminal tail (CT) in binding, signaling, and receptor internalization. Receptors truncated at amino acids 425, 418, and 405, expr essed in COS-7 or CHO-K1 cells, exhibited similar binding to wild type rece ptors, GIP-dependent cAMP production with the 405 mutant was decreased in C OS-7 cells. Maximal cAMP production in CHO-K1 cells was reduced with all tr uncated forms. Binding was undetectable with a receptor truncated at amino acid 400; increasing tail length by adding 5 alanines restored binding and signaling. Mutants produced by alanine scanning of residues 394-401, adjace nt to transmembrane domain 7, were all functional. CT truncation by 30 or m ore amino acids, mutation of serines 426/427, singly or combined, or comple te CT serine knockout all reduced receptor internalization rate, The majori ty of the GIP receptor CT is therefore not required for signaling, a minimu m chain length of similar to 405 amino acids is needed for receptor express ion, and serines 426 and 427 are important for regulating rate of receptor internalization.