The N-terminal thrombin receptor fragment SFLLRN, but not catalytically inactive thrombin-derived agonists, activate U937 human monocytic cells: evidence for receptor hydrolysis in thrombin-dependent signalling.
It has previously been reported that murine macrophages can respond chemotactically and mitogenically to the serine proteinase thrombin. There is a similar response in these macrophages to catalytically inactivated thrombin or to peptide fragments of the thrombin B-chain [Bar-Shavit, Kahn, Mann and Wilner (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 976-980]. However, the existence of a non-proteolytic mechanism of thrombin receptor activation in mononuclear cells was not evident in the present study using U937 human monocytic cells. The ability of thrombin to stimulate intracellular Ca2+ mobilization, actin polymerization or cell proliferation was not mimicked by N alpha-tosyl-L-lysine chloromethyl ketone (TLCK)-treated thrombin or by a synthetic 14-amino-acid peptide (single amino acid letter code YPPWNKNFTENDLL) corresponding to a part of the B-chain of thrombin which was reported to be mitogenic in murine macrophages. Evidence was obtained, however, in U937 cells for the presence of proteolytic-dependent thrombin receptor similar to the thrombin receptor expressed in platelets, which following thrombin cleavage exposes a new N-terminal tethered ligand. In support of this, a thrombin-receptor-derived hexapeptide (TRP; sequence SFLLRN), corresponding to a part of the thrombin receptor tethered ligand, mimicked all the actions of thrombin in U937 cells. Further, TRP and thrombin cross-desensitized U937 cells to subsequent stimulation with either TRP or thrombin, suggesting that TRP acted through the same U937 cell surface receptor as did thrombin. Thrombin activation of U937 monocytic cells can therefore be accounted for entirely by a proteolytic mechanism of thrombin receptor activation.
Department of Clinical Pharmacology, Royal Postgraduate Medical School, London, U.K.
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Joseph
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MacDermot
D000199:Actins / Q000096:biosynthesis; D000595:Amino Acid Sequence; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D018179:Receptors, Thrombin; D015398:Signal Transduction; D013917:Thrombin / Q000378:metabolism* / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D014107:Tosyllysine Chloromethyl Ketone / Q000031:analogs & derivatives / Q000494:pharmacology
D000199:Actins; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; D014107:Tosyllysine Chloromethyl Ketone; D013917:Thrombin; C050816:tosyllysine chloromethyl ketone-thrombin; D002118:Calcium; D013936:Thymidine