Design of short-sequenced collagen type III-derived peptides with GPR56 dependent biological activity

We have previously shown that activation of G protein-coupled receptor 56 (GPR56) by its endogenous ligand collagen type III (Coll III) improves pancreatic β-cell function and potentiates glucose-stimulated insulin secretion (GSIS). In the current study, alignment of Coll III and GPR56 genomic DNA revealed three complementary nucleotide regions, enabling the design of three short peptides (NQGCKL.NH2, H.LLSSR.NH2 and NH2.GYYCG.COOH). The peptides in double or triple combinations markedly increased cellular cAMP levels, enhanced GSIS, and improved cell viability both in human (EndoC βH1 cells) and rat pancreatic β-cell lines (INS-1 832/13 cells). These effects were markedly absent in GPR56-knockdown (GPR56-KD) cells, supporting receptor-dependent specificity. The peptide mixtures also counteracted cytokine- and glucotoxicity-induced cellular stress, restoring cAMP generation and insulin secretion. Comparable effects were not observed with dapagliflozin while glibenclamide decreased cell viability under similar conditions. In hepatic cell lines (HepG2, Hep1-6), peptide combinations enhanced cell survival in a GPR56-dependent manner, consistent with a broader role of GPR56 in metabolic regulation. Collectively, the data identify three short sequence collagen III-derived peptides whose biological effects are dependent on GPR56 expression and support their further investigation as tools for studying GPR56-associated cellular signaling for the future in vivo validation.