Metalloproteinase-mediated regulation of L-selectin levels on leucocytes.
Leucocyte (L)-selectin can be proteolytically cleaved in the membrane proximal extracellular region to yield a soluble fragment that contains the functional lectin and epidermal growth factor domains. A variety of stimuli are known to stimulate L-selectin shedding including chemoattractants, phorbol esters, and L-selectin cross-linking; however, the enzymes that regulate L-selectin expression are not characterized. In this study we have used phorbol ester to stimulate endoproteolytic release of L-selectin and identified a major role for a cell surface metalloproteinase (L-selectin sheddase) in this process. The hydroxamic acid-based inhibitor of zinc-dependent matrix metalloproteinases Ro 31-9790 completely prevented shedding of cell surface L-selectin from leucocytes in mouse, rat, and man. L-selectin was susceptible to cleavage by known matrix metalloproteinases. Recombinant human fibroblast collagenase (MMP1) reduced the number of L-selectin-positive lymphocytes to a similar extent as phorbol ester activation, and stromelysin (MMP3) had a partial effect on L-selectin expression. Gelatinases A (MMP2) and B (MMP9) were without effect. Lymphocytes did not express fibroblast collagenase or stromelysin at the cell surface, and tissue inhibitor of metalloproteinases (TIMP) did not affect L-selectin levels. L-selectin sheddase was not detected in media harvested from phorbol ester-stimulated lymphocytes and was only able to cleave L-selectin in the cis but not the trans configuration. These results suggest that endoproteolytic release of L-selectin from the leucocyte surface is mediated by a metalloproteinase (L-selectin sheddase), which is distinguishable from known matrix metalloproteinases. Understanding the regulation of L-selectin sheddase will be critical for controlling leucocyte migration from the blood.
Preece
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G
Division of Cellular Immunology, National Institute for Medical Research, London, United Kingdom.
Murphy
G
G
Ager
A
A
D000818:Animals; D002918:Chymotrypsin / Q000494:pharmacology; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D019041:L-Selectin / Q000032:analysis*; D007962:Leukocytes / Q000737:chemistry*; D008666:Metalloendopeptidases / Q000502:physiology*; D051379:Mice; D008807:Mice, Inbred BALB C; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D051381:Rats
D006877:Hydroxamic Acids; D011480:Protease Inhibitors; C097661:Ro 31-9790; D019041:L-Selectin; D015240:Phorbol 12,13-Dibutyrate; D002918:Chymotrypsin; D008666:Metalloendopeptidases