Macrophage migration inhibitory factor (MIF) but not its homologue D‐dopachrome tautomerase (D‐DT) promotes fibroblast motility in a CD44/CD74‐independent manner

Abstract

Macrophage migration inhibitory factor (MIF) and D‐dopachrome tautomerase (D‐DT) are paralogous pro‐inflammatory cytokines with chemokine‐like functions. Whereas cytosolic MIF plays a role in cell cycle, secreted MIF and D‐DT activate second messenger signalling through a CD44/CD74 receptor complex. To differentiate between receptor‐ and non‐receptor‐mediated events, COS‐7/M6 fibroblasts deficient in CD44/CD74 (WT) were employed to generate stable cell lines expressing CD44 and CD74. Chemokinesis (random single‐cell motility) was assessed in cells stimulated with native recombinant MIF/D‐DT, enzyme‐null recombinant MIF, inhibitors of MIF's tautomerase activity, and inhibitors of clathrin‐ and non‐clathrin‐dependent endocytosis. In the presence of CD44/CD74, both MIF and D‐DT stimulated chemokinesis but only MIF enhanced chemokinesis in the absence of receptors. The stimulatory effect of MIF on migration depended on its tautomerase activity and lipid raft/caveolae‐mediated, but not clathrin‐mediated endocytosis. To observe a direct effect of MIF and D‐DT on actin dynamics, actin polymerization was quantified in an in vitro actin assembly assay. MIF but not D‐DT decreased the rate of F‐actin assembly. By decreasing the rate of cell extract‐driven actin assembly, MIF phenocopies the function of F‐actin capping proteins. Moreover, stimulation with MIF increased the number of prominent F‐actin stress fibres in COS‐7/M6 WT. Taken together, MIF stimulates fibroblast chemokinesis independent of its receptors, likely by directly modulating the actin cytoskeleton. MIF‐mediated chemokinesis appears to depend on lipid raft/caveolae‐mediated endocytosis. D‐DT triggers chemokinesis only in the presence of MIF/D‐DT receptors – CD44/CD74 – implying D‐DT requires the classical receptor‐driven second‐messenger transduction pathway.Support or Funding InformationIntramural funding.