Monomeric midkine induces tumor cell proliferation in the absence of cell-surface proteoglycan binding

Midkine (MK), a retinoic acid-inducible heparin-binding protein, is a mitog en which initiates a cascade of intracellular protein tyrosine phosphorylat ion mediated by the JAK/STAT pathway after binding to its high affinity p20 0(+)/MKR cell surface receptor in the G401 cell line [Ratovitski, E. A. (19 98) J. Biol. Chem, 273, 3654-3660]. In this study, we determined the biophy sical characteristics of purified recombinant murine MK and analyzed the re quirements for Ligand multimerization and cell surface proteoglycan binding for the G401 cell mitogenic activity of MK. Our studies indicate that the secreted form of MK (M = 13 kDa) exists in solution as an asymmetric monome r with a frictional coefficient of 1.48 and a Stokes radius of 23.7 Angstro m. By constructing bead models of MK using the program AtoB and the program HYDRO to predict the hydrodynamic properties of each model, our data sugge st that MK has a dumb-bell shape in solution composed of independent N- and C-terminal domains separated by an extended linker. This asymmetric MK mon omer is a biologically active ligand with mitogenic activity on G401 cells in vitro. Neither heparin-induced formation of noncovalent MK multimers nor tissue transglutaminase II covalent multimerization of MK enhanced MK mito genic activity in this system. Since neither heparin competition nor cell t reatment with chondroitinase ABC or heparinase III abolished the mitogenic effects of MK on G401 cells, cell-surface proteoglycan binding by MK does n ot appear to be a requirement for its observed mitogenic effects. These res ults provide strong evidence that the MK-specific p200(+)/MKR has distincti ve biochemical properties which distinguish it from the receptor tyrosine p hosphatase cell-surface proteoglycan PTPR zeta/PTP beta and support the hyp othesis that the diverse biological effects of MK are mediated by multiple cell-specific signal transduction receptors.