IDENTIFICATION OF A HEPATOCYTE GROWTH-FACTOR AUTOCRINE LOOP IN A MURINE MAMMARY-CARCINOMA

Constitutive activation of growth factor receptors through autocrine/p aracrine mechanisms occurs frequently in human cancers and is thought to play an important role in carcinogenesis, We have demonstrated prev iously that hepatocyte growth factor (HGF) is a potent mitogenic facto r for murine mammary carcinoma (SP1) cells in vitro, We report here an autocrine HGF loop in SP1 cells. HGF receptor/Met is expressed in SP1 cells and is constitutively tyrosine phosphorylated, The phosphorylat ion of HGF receptor/ Met is inhibited when cells are exposed to surami n or anti-HGF IgG, This finding suggests that constitutive tyrosine ph osphorylation of HGF receptor/Met is sustained by an extracellular fac tor, most likely HGF, Using Northern blot and Western blot analysis, w e detected expression of a 6-kb HGF mRNA in SP1 cells and a M(r) 85,00 0 HGF protein in SP1-conditioned medium, respectively, In vitro transl ation of mRNA from SP1 cells and metabolic labeling confirmed expressi on and synthesis of HGF by SP1 cells, SP1 cells also invade through Ma trigel-coated transwell membranes in an in vitro invasion assay, and i nvasion of these cells was inhibited by neutralizing anti-HGF IgG, In addition, SP1-conditioned medium induced scatter activity of Madin-Dar by canine kidney epithelial cells, and this activity was inhibited by neutralizing anti-HGF IgG, We have also shown that several signaling m olecules including phosphatidylinositol 3-kinase, Src, focal adhesion kinase, and phospholipase C-gamma in SP1 cells are constitutively tyro sine phosphorylated, suggesting that coexpression of HGF and HGF recep tor/Met may in part contribute to sustained tyrosine phosphorylation o f these cytoplasmic proteins in SP1 cells. Our observations in the SP1 model suggest that HGF contributes to growth and invasive phenotypes of mammary carcinomas via both paracrine and autocrine mechanisms.