Subcellular distribution and cytokine- and chemokine-regulated secretion of leukolysin/MT6-MMP/MMP-25 in neutrophils.
Leukolysin, originally isolated from human leukocytes, is the sixth member of the membrane-type matrix metalloproteinase (MT-MMP) subfamily with a potential glycosylphosphatidylinositol (GPI) anchor. To understand its biological functions, we screened subpopulations of leukocytes and localized the expression of leukolysin at the mRNA level to neutrophils. Polyclonal and mono-specific antisera raised against a synthetic peptide from its hinge region recognized a major protein species at 56 kDa and several minor forms between 38 and 45 kDa in neutrophil lysates. In resting neutrophils, leukolysin is distributed among specific granules ( approximately 10%), gelatinase granules ( approximately 40%), secretory vesicles ( approximately 30%), and the plasma membrane ( approximately 20%), a pattern distinct from that of neutrophil MMP-8 and MMP-9. Consistent with its membrane localization and its reported GPI anchor, leukolysin partitions into the detergent phase of Triton X-114 and can be released from intact resting neutrophils by glycosylphosphatidylinositol-specific phospholipase C. Phorbol myristate acetate stimulates neutrophils to discharge 100% of leukolysin from specific and gelatinase granules and approximately 50% from the secretory vesicles and plasma membrane, suggesting that leukolysin can be mobilized by physiological signals to the extracellular milieu as a soluble enzyme. Indeed, interleukin 8, a neutrophil chemoattractant, triggered a release of approximately 85% of cellular leukolysins by a process resistant to a mixture of proteinase inhibitors, including aprotinin, BB-94, pepstatin, and E64. Finally, purified recombinant leukolysin can degrade components of the extracellular matrix. These results not only establish leukolysin as the first neutrophil-specific MT-MMP but also implicate it as a cytokine/chemokine-regulated effector during innate immune responses or tissue injury.
Departments of Pharmacology and Neurosciences, University of Minnesota School of Medicine, Minneapolis, Minnesota 55455, USA.
T
T
Kang
J
J
Yi
A
A
Guo
X
X
Wang
C M
CM
Overall
W
W
Jiang
R
R
Elde
N
N
Borregaard
D
D
Pei
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D016207:Cytokines / Q000494:pharmacology*; D003594:Cytoplasmic Granules / Q000201:enzymology; D004285:Dogs; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007375:Interleukin-1 / Q000494:pharmacology; D016209:Interleukin-8 / Q000494:pharmacology; D020784:Matrix Metalloproteinase 8 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D053504:Matrix Metalloproteinases, Membrane-Associated; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D011994:Recombinant Proteins / Q000032:analysis / Q000378:metabolism; D013347:Subcellular Fractions / Q000201:enzymology; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
D016207:Cytokines; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols; D007375:Interleukin-1; D016209:Interleukin-8; D011994:Recombinant Proteins; D010738:Type C Phospholipases; D020782:Matrix Metalloproteinases; D053504:Matrix Metalloproteinases, Membrane-Associated; C403928:matrix metalloproteinase 25; D020784:Matrix Metalloproteinase 8; D020780:Matrix Metalloproteinase 9; D043265:Phosphatidylinositol Diacylglycerol-Lyase