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Identification and molecular cloning of a novel mouse mucosal mast cell serine protease
W E Serafin1, D S Reynolds, S Rogelj
1Department of Medicine, Harvard Medical School, Boston, Massachusetts.
Abstract:
A novel 28,000 Mr serine protease, designated mouse mast cell protease-2 (MMCP-2), that is stored in the secretory granules of Kirsten sarcoma virus-immortalized mouse mast cells (KiSV-MC) has been identified and its NH2-terminal amino acid sequence has been determined. Analysis of a 953-base pair cDNA that encodes MMCP-2 revealed that this serine protease is a basically charged protein, possessing the histidine-aspartic acid-serine charge relay system that is characteristic of other serine proteases. DNA blot analysis using the full-length MMCP-2 cDNA indicated the existence of a family of highly related serine protease genes in the mouse genome. When the same DNA blot was probed with the 149-base pair KpnI----3' fragment of the cDNA, the probe hybridized to a single DNA fragment, thereby demonstrating that this 3' fragment could be used as a gene-specific probe. The presence of high levels of the MMCP-2 mRNA transcript in the intestines of nematode-infected mice, and its absence in mouse bone marrow-derived mast cells and peritoneal cavity-derived connective tissue mast cells, suggest that this member of the mouse mast cell protease family is preferentially expressed late in the differentiation of mucosal mast cells.
Insights
Researchers identified mouse mast cell protease-2 (MMCP-2), a novel serine protease. MMCP-2 is preferentially expressed late in mucosal mast cell differentiation, particularly in nematode-infected mice.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Mast cells are key immune cells involved in allergic reactions and host defense.
- Serine proteases are a class of enzymes with diverse biological functions.
- Understanding mast cell proteases is crucial for deciphering immune responses.
Purpose of the Study:
- To identify and characterize a novel serine protease from mouse mast cells.
- To determine the gene structure and expression patterns of this new protease.
- To investigate its potential role in mast cell differentiation and immune responses.
Main Methods:
- Purification and sequencing of the novel serine protease (MMCP-2).
- cDNA cloning and sequence analysis of MMCP-2.
- DNA blot analysis to study gene family and specificity.
- Northern blot analysis to examine mRNA expression in different mast cell types and tissues.
Main Results:
- Identification of a novel 28,000 Mr serine protease, MMCP-2, in Kirsten sarcoma virus-immortalized mouse mast cells.
- MMCP-2 is a basic serine protease with a characteristic catalytic triad.
- DNA blot analysis revealed a family of related serine protease genes; a 3' fragment of MMCP-2 cDNA served as a gene-specific probe.
- MMCP-2 mRNA was highly expressed in the intestines of nematode-infected mice but absent in bone marrow-derived mast cells and peritoneal connective tissue mast cells.
Conclusions:
- MMCP-2 is a distinct member of the mouse mast cell protease family.
- Its expression pattern suggests preferential late-stage differentiation in mucosal mast cells.
- MMCP-2 may play a specific role in mucosal immunity, particularly during parasitic infections.