title: Phorbol ester activation of a proteolytic cascade capable of activating latent transforming growth factor-betaL a process initiated by the exocytosis of cathepsin B.
abstract: 12-O-Tetradecanoylphorbol-13-acetate (TPA) suppresses the proliferation of the human breast epithelial cell line MCF10A-Neo by initiating proteolytic processes that activate latent transforming growth factor (TGF)-beta in the serum used to supplement culture medium. Within 1 h of treatment, cultures accumulated an extracellular activity capable of cleaving a substrate for urokinase-type plasminogen activator (uPA) and tissue plasminogen activator (tPA). This activity was inhibited by plasminogen activator inhibitor-1 or antibodies to uPA but not tPA. Pro-uPA activation was preceded by dramatic changes in lysosome trafficking and the extracellular appearance of cathepsin B and beta-hexosaminidase but not cathepsins D or L. Co-treatment of cultures with the cathepsin B inhibitors CA-074 or Z-FA-FMK suppressed the cytostatic effects of TPA and activation of pro-uPA. In the absence of TPA, exogenously added cathepsin B activated pro-uPA and suppressed MCF10A-Neo proliferation. The cytostatic effects of both TPA and cathepsin B were suppressed in cells cultured in medium depleted of plasminogen/plasmin or supplemented with neutralizing TGF-beta antibody. Pretreatment with cycloheximide did not suppress the exocytosis of cathepsin B or the activation of pro-uPA. Hence, TPA activates signaling processes that trigger the exocytosis of a subpopulation of lysosomes/endosomes containing cathepsin B. Subsequently, extracellular cathepsin B initiates a proteolytic cascade involving uPA, plasminogen, and plasmin that activates serum-derived latent TGF-beta.
authors.affiliation: Institute of Environmental Health Sciences, Wayne State University and the Department of Pharmacology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
authors.forename: Meng
authors.initials: M
authors.lastname: Guo
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Mathieu
authors.forename: Bruce
authors.initials: B
authors.lastname: Linebaugh
authors.forename: Bonnie F
authors.initials: BF
authors.lastname: Sloane
authors.forename: John J
authors.initials: JJ
authors.lastname: Reiners
mesh_terms: D002384:Catalysis; D002401:Cathepsin B / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D004789:Enzyme Activation; D005089:Exocytosis; D005341:Fibrinolysin / Q000502:physiology; D006801:Humans; D006868:Hydrolysis; D010958:Plasminogen / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
chemical_list: D016212:Transforming Growth Factor beta; D010958:Plasminogen; D003513:Cycloheximide; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator; D002401:Cathepsin B; D013755:Tetradecanoylphorbol Acetate