Identification of a protein hydrolysate responsive G protein-coupled receptor in enterocytes

Abstract

G protein-coupled receptors (GPCRs) have the potential to play a role as molecular sensors responsive to luminal dietary contents. Although such a role for GPCRs has been implicated in the intestinal response to protein hydrolysate, no GPCR directly involved in this process has been previously identified. In the present study, for the first time, we identified GPR93 expression in enterocytes and demonstrated its activation in these cells by protein hydrolysate with EC50 of 10.6 mg/ml as determined by the induction of intracellular free Ca2+. In enterocytes, GPR93 was synergistically activated by protein hydrolysate in combination with an agonist, oleoyl-l-α-lysophosphatidic acid (LPA), which activated the receptor in these enterocytes with EC50 of 7.9 nM. The increased intracellular Ca2+ by GPR93 activation was observed without the addition of a promiscuous Gα protein and was pertussis toxin sensitive, which suggests Gαq- and Gαi-mediated pathways. Activated GPR93 also induced pertussis toxin-sensitive ERK1/2 phosphorylation. Both nuclear factor of activated T cells and 12- O-tetradecanoylphorbol 13-acetate responsive elements reporter activities were induced by protein hydrolysate in cells exogenously expressing GPR93. The peptidomimetic cefaclor by itself did not activate GPR93 but potentiated the protein hydrolysate response and further amplified the synergistic enhancement of GPR93 activation by protein hydrolysate and LPA. These data suggest that, physiologically, the composition of stimuli might determine GPR93 activity or its sensitivity toward a given activator and suggest a new mechanism of the regulation of mucosal cell proliferation and differentiation and hormonal secretion by dietary products in the lumen.

Soybean β-Conglycinin Bromelain Hydrolysate Stimulates Cholecystokinin Secretion by Enteroendocrine STC-1 Cells to Suppress the Appetite of Rats under Meal-Feeding Conditions

“…Recent reports have shown that a G-protein coupled receptor (GPCR), GPR93, mediated the peptone-induced CCK secretion in STC-1 cells, 26,27) and we have demonstrated the involvement of the Gq pathway for BconP-induced CCK secretion in these cells. [28][29][30] We have recently found the calcium-sensing receptor, CaR, to function as the receptor for the arginine-rich 51-63 peptide in -conglycinin by inducing intracellular Ca 2þ mobilization for triggering subsequent CCK secretion in EECs.…”

Feeding-dependent activation of enteric cells and sensory neurons by lymphatic fluid: evidence for a neurolymphocrine system

“…The closest structurally related GPCRs to LPA5 are P2Y5 and P2Y9, which are also activated by LPA (16,46). However, we found that only LPA5 is activated by partially digested dietary protein (peptone).…”

“…The mitochondria-targeted aequorin (mtAEQ) and LPA5 expression vectors were constructed as previously described (16). The vector base for the reporter constructs, pBVluc, was a generous gift from Dr. Bert Vogelstein (The Johns Hopkins Kimmel Cancer Center).…”

“…The GPCR lysophosphatidic acid 5 (LPA5) (also called GPR93 or GPR92) is expressed in mucosal cells (16) and neuronal tissues, such as the spinal cord dorsal horns (50) and dorsal root ganglia (DRG) (59). The closest structurally related GPCRs to LPA5 are P2Y5 and P2Y9, which are also activated by LPA (16,46).…”

“…LPA5 responds to both LPA and peptone (with EC 50 of 8 nM and 10 mg/ml, respectively), yielding distinct effector pathways (17). Activation of LPA5 by peptone in the EEC STC1 cell line induces a calcium-dependent CCK release and synthesis, whereas LPA activation does not result in the release of CCK (16). This allows for a regulation of CCK release, by the presence of partially digested dietary protein that is independent of the presence of LPA.…”