Protein kinase C-dependent in vivo phosphorylation of prourokinase leads to the formation of a receptor competitive antagonist.
We recently reported that in vivo phosphorylation of urokinase-type plasminogen activator on Ser138/303 prevents its catalytic-independent ability to promote myelomonocytic cell adherence and motility. We now show that Ca2+ activated, phospholipid-dependent protein kinase C from rat brain phosphorylates in vitro a peptide corresponding to prourokinase residues 133-143 (DGKKPSSPPEE) and the full-length molecule on Ser138/139. The in vivo involvement of the protein kinase C isoenzyme family is supported by the finding that inhibition of kinase C activity prevents prourokinase phosphorylation on Ser138/303 in A431 human carcinoma cells. Conversely, a short treatment of A431 cells with phorbol myristate acetate increases the extent of phosphorylated prourokinase and, concomitantly, affects its function; under these conditions, the capability of prourokinase to up-regulate U937 monocyte-like cell adherence is severely impaired, although receptor binding is unaltered. By the aid of a "phosphorylation-like" variant (Ser138 to Glu) we show that modification of Ser138 is sufficient to confer to prourokinase the antagonistic properties observed following in vivo stimulation of protein kinase C activity. These observations provide the first evidence that protein kinase C directs the formation of a receptor competitive antagonist by regulating the in vivo phosphorylation state of prourokinase.
International Institute of Genetics and Biophysics, C.N.R., 80125 Naples, Italy.
P
P
Franco
O
O
Massa
M
M
Garcia-Rocha
F
F
Chiaradonna
C
C
Iaccarino
I
I
Correas
E
E
Mendez
J
J
Avila
F
F
Blasi
M P
MP
Stoppelli
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology; D002448:Cell Adhesion; D015536:Down-Regulation; D004792:Enzyme Precursors / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors*; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D020298:U937 Cells; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D004792:Enzyme Precursors; D007527:Isoenzymes; D009842:Oligopeptides; C524898:PLAUR protein, human; C524900:Plaur protein, rat; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins; D012694:Serine; D011493:Protein Kinase C; D014568:Urokinase-Type Plasminogen Activator; C067551:saruplase