Distinct pathways of cell migration and antiapoptotic response to epithelial injury: Structure-function analysis of human intestinal trefoil factor

The trefoil peptide intestinal trefoil factor (ITF) plays a critical role i n the protection of colonic mucosa and is essential to restitution after ep ithelial damage. These functional properties are accomplished through coord inated promotion of cell migration and inhibition of apoptosis. ITF contain s a unique three-looped trefoil motif formed by intrachain disulfide bonds among six conserved cysteine residues, which is thought to contribute to it s marked protease resistance. ITF also has a seventh cysteine residue, whic h permits homodimer formation. A series of cysteine-to-serine substitutions and a C-terminally truncated ITF were made by PCR site-directed mutagenesi s. Any alteration of the trefoil motif or truncation resulted in loss of pr otease resistance. However, neither an intact trefoil domain nor dimerizati on was required to promote cell migration. This pro-restitution activity co rrelated with the ability of the ITF mutants to activate mitogen-activated protein (MAP) kinase independent of phosphorylation of the epidermal growth factor (EGF) receptor. In contrast, only intact ITF retained both phosphat idylinositol 3-kinase and the EGF receptor-dependent antiapoptotic effect i n HCT116 and IEC-6 cells. The inability to block apoptosis correlated with a loss of trefoil peptide-induced transactivation of the EGF receptor or Ak t kinase in HT-29 cells. In addition to defining structural requirements fo r the functional properties of ITF, these findings demonstrate that distinc t intracellular signaling pathways mediate the effects of ITF on cell migra tion and apoptosis.