title: Phorbol 12-myristate 13-acetate-induced ectodomain shedding and phosphorylation of the human meprinbeta metalloprotease.
abstract: Shedding of proteins localized at the cell surface is an important regulatory step in the function of many of these proteins. Human meprin (N-benzoyl-l-tyrosyl-p-aminobenzoic acid hydrolase, PPH, EC 3.4.24.18) a zinc-metalloendopeptidase of the astacin family is an oligomeric protein complex of alpha- and beta-subunits and is expressed abundantly in the intestine and kidney as well as in leukocytes of the lamina propria and in cancer cells. In transfected cells intracellular proteolytic removal of the membrane anchor results in the secretion of the meprin alpha-subunit. In rats and mice, the beta-subunit exists in a membrane-anchored form. In contrast, human meprinbeta is constitutively converted into a secretable form. We now show that phorbol 12-myristate 13-acetate (PMA) stimulates an increased release of hmeprinbeta from transfected COS-1 cells, whereas hmeprinalpha secretion is not influenced. This stimulatory effect is inhibited by the protein kinase C (PKC) inhibitor staurosporine, suggesting that activation of PKC mediates PMA-induced hmeprinbeta shedding. The use of different protease inhibitors shows that two different metalloprotease activities are responsible for the constitutive and the PMA-stimulated hmeprinbeta shedding. We identified tumor necrosis factor alpha-converting enzyme (TACE or ADAM17) as the protease that mediates the PMA-induced release. We also demonstrate that hmeprinbeta is phosphorylated by PMA treatment on Ser687 within a PKC consensus sequence in the cytosolic domain of the protein. This phosphorylation of hmeprinbeta is not, however, implicated in the enhanced secretion by PMA treatment.
authors.affiliation: Institute of Biochemistry and Molecular Biology and Department of Pediatrics, Berne University, 3012 Berne, Switzerland.
authors.forename: Dagmar
authors.initials: D
authors.lastname: Hahn
authors.forename: Anastassios
authors.initials: A
authors.lastname: Pischitzis
authors.forename: Sandra
authors.initials: S
authors.lastname: Roesmann
authors.forename: Marianne K
authors.initials: MK
authors.lastname: Hansen
authors.forename: Boris
authors.initials: B
authors.lastname: Leuenberger
authors.forename: Ursula
authors.initials: U
authors.lastname: Luginbuehl
authors.forename: Erwin E
authors.initials: EE
authors.lastname: Sterchi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D008954:Models, Biological; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011233:Precipitin Tests; D011480:Protease Inhibitors / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D019311:Staurosporine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate; D013997:Time Factors; D014162:Transfection
chemical_list: D004791:Enzyme Inhibitors; D011480:Protease Inhibitors; D011493:Protein Kinase C; D008666:Metalloendopeptidases; C095663:meprin B; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate