Identification of human cysteine-rich secretory protein 3 (CRISP-3) as a matrix protein in a subset of peroxidase-negative granules of neutrophils and in the granules of eosinophils.
Cysteine-rich secretory protein 3 (CRISP-3; also known as SGP28) was originally discovered in human neutrophilic granulocytes. We have recently developed a sensitive sandwich enzyme-linked immunosorbent assay for CRISP-3 and demonstrated the presence of CRISP-3 in exocrine secretions. To investigate the subcellular localization and mobilization of CRISP-3 in human neutrophils, we performed subcellular fractionation of resting and activated neutrophils on three-layer Percoll density gradients, release-studies of granule proteins in response to different secretagogues, and double-labeling immunogold electron microscopy. CRISP-3 was found to be localized in a subset of granules with overlapping characteristics of specific and gelatinase granules and mobilized accordingly, thus confirming the hypothesis that peroxidase-negative granules exist as a continuum from specific to gelatinase granules regarding protein content and mobilization. CRISP-3 was found to be a matrix protein, which is stored in granules as glycosylated and as unglycosylated protein. The subcellular distribution of the two forms of CRISP-3 was identical. In addition, CRISP-3 was found as a granule protein in eosinophilic granulocytes. The presence of CRISP-3 in peroxidase-negative granules of neutrophils, in granules of eosinophils, and in exocrine secretions indicates a role in the innate host defense.
Udby
Lene
L
Granulocyte Research Laboratory, Department of Hematology, Rigshospitalet, Copenhagen, Denmark. l.udby@rh.dk
Calafat
Jero
J
Sørensen
Ole E
OE
Borregaard
Niels
N
Kjeldsen
Lars
L
D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D003594:Cytoplasmic Granules / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D047209:Eosinophil Peroxidase; D004804:Eosinophils / Q000737:chemistry* / Q000187:drug effects / Q000648:ultrastructure; D018093:Gelatinases / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007781:Lactoferrin / Q000378:metabolism; D016253:Microscopy, Immunoelectron; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009195:Peroxidase / Q000378:metabolism; D010544:Peroxidases / Q000032:analysis; D011499:Protein Processing, Post-Translational; D012471:Salivary Proteins and Peptides / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D029589:Seminal Plasma Proteins / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D013347:Subcellular Fractions / Q000737:chemistry; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015054:Zymosan / Q000494:pharmacology
C489926:CRISP3 protein, human; D012471:Salivary Proteins and Peptides; D029589:Seminal Plasma Proteins; D015759:Ionomycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015054:Zymosan; D047209:Eosinophil Peroxidase; D010544:Peroxidases; D009195:Peroxidase; D007781:Lactoferrin; D018093:Gelatinases; D013755:Tetradecanoylphorbol Acetate