Proinsulin C-peptide and its analogues induce intracellular Ca2+ increases in human renal tubular cells

Based on the findings that proinsulin C-peptide binds specifically to cell membranes, we investigated the effects of C-peptide and related molecules on the intracellular Ca2+ concentration ([Ca2+](i)) in human renal tubular cells using the indicator fura-2/AM. The results show that human C-peptide and its C-terminal pentapeptide (positions 27-31, EGSLQ), but not the des (27-31) C-peptide or randomly scrambled C- peptide, elicit a transient increase in [Cal(2+)](i). Rat C-peptide and rat C-terminal pentapeptide also induce a [Ca2+](i) response in human tubular cells, while a human pentapeptide analogue with Ala at position 1 gives no [Ca2+](i) response, and those with Ala at positions 2-5 induce responses with different amplitudes. These results define a species cross-reactivity for C-peptide and demonstrate the importance of Glu at position 1 of the pentapeptide. Preincubation of cells with pertussis toxin abolishes the effect on [Ca2+](i) by both C-peptide and the pentapeptide. These results are compatible with previous data on C-peptide binding to cells and activation of Na+,K(+)ATPase. Combined, all data show that C-peptide is a bioactive peptide and suggest that it elicits changes in [Ca2+](i) via G-protein-coupled pathways, giving downstream enzyme effects.