Calmodulin inhibitors trigger the proteolytic processing of membrane type-1 matrix metalloproteinase, but not its shedding in glioblastoma cells

Most transmembrane proteins are subjected to limited proteolysis by cellula r proteases, and stimulation of cleavage of membrane proteins by calmodulin (CaM) inhibitors was recently shown. The present study investigated the ab ility of several CaM inhibitors to induce the proteolytic cleavage of the m embrane type-1 matrix metalloproteinase (MT1-MMP) from the cell surface of highly invasive U-87 glioblastoma cells. Although no shedding of a soluble MT1-MMP form was induced by CaM inhibitors in the conditioned media, we sho wed that these inhibitors induced MT1-MMP proteolytic processing to the 43 kDa membrane-bound inactive form that was not correlated with an increase i n proMMP-2 activation but rather with an increase in tissue inhibitor of MM Ps (TIMP)-2 expression levels. Moreover, this proteolytic processing was se nsitive to marimastat suggesting the involvement of MMPS. Interestingly, Ca M inhibitors antagonized concanavalin A- and cytochalasin D-induced proMMP- 2 activation, and affected the cytoskeletal actin organization resulting in the loss of migratory potential of U-87 glioblastoma cells. Cytoplasmic ta il-truncated MT1-MMP constructs expressed in COS-7 cells were also affected by CaM inhibitors suggesting that these inhibitors stimulated MT1-MMP prot eolytic processing by mechanisms independent of the CaM-substrate interacti on. We also propose that TIMP-2 acts as a negative regulator of MTI-MMP-dep endent activities promoted by the action of CaM inhibitors in U-87 glioblas toma cells.