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Expression profile of novel members of the rat mast cell protease (rMCP)-2 and (rMCP)-8 families, and functional analyses of mouse mast cell protease (mMCP)-8
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
2007 (English)In: Immunogenetics, ISSN 0093-7711, E-ISSN 1432-1211, Vol. 59, no 5, 391-405 p.Article in journal (Refereed) Published
Abstract [en]

Four hematopoietic serine proteases are common to the mast cell chymase locus of all analyzed mammals: α-chymase, cathepsin G, granzyme B, and granzyme C/H. Apart from these common genes, the mouse and rat loci hold additional granzyme-, β-chymase-, and Mcpt8-like genes. To better understand the functional consequences of these additional enzymes and to be able to compare human and rodent immune functions, we have analyzed the expression of novel β-chymase- and Mcpt8-like genes in the rat. Four novel genes, i.e., Mcpt2-rs2a, Mcpt2-rs2c, Mcpt8-rs1, and Mcpt8-rs4 were transcribed in tissues holding mucosal mast cells (MMC), where also the classical MMC protease Mcpt2 was expressed. We also found transcripts of rat vascular chymase (rVch) in some of these tissues. RVch is a β-chymase that converts angiotensin I, like the human chymase. Rat MMC may therefore have similar angiotensin-converting properties as chymase-positive human mast cells, although these are mostly regarded the counterpart of rat connective tissue mast cells. The human mast cells that are considered the counterpart of rat MMC express, however, only tryptase, whereas rat MMC express various proteases, but no tryptase. We further studied the proteolytic activity of mMCP-8 as a first representative for the Mcpt8-subfamily. Based on sequence comparison and molecular modeling, mMCP-8 may prefer aspartic acid in substrate P1 position. However, we could not detect hydrolysis of chromogenic substrates or phage-displayed random nonapeptides despite numerous trials. On the other hand, we have obtained evidence that the function of the Mcpt8-like proteases depends on proteolytic activity. Namely, the expression of the only Mcpt8-family member with a mutation in the catalytic triad, Mcpt8-rs3, was strongly reduced. Thus, the substrate specificity of mMCP-8 may be too narrow to be detected with the employed methods, or the enzyme may require a substrate conformation that is not provided by the analyzed peptides.

Place, publisher, year, edition, pages
2007. Vol. 59, no 5, 391-405 p.
Keyword [en]
Angiotensin, Chymase locus, Gene expression, Mast cell, mMCP-8, rMCP-2, rMCP-8, Serine protease
National Category
Biological Sciences
URN: urn:nbn:se:uu:diva-95244DOI: 10.1007/s00251-007-0202-1ISI: 000245429400006PubMedID: 17342483OAI: oai:DiVA.org:uu-95244DiVA: diva2:169382
Available from: 2006-11-29 Created: 2006-11-29 Last updated: 2011-02-08Bibliographically approved
In thesis
1. Sculpted through Time: Evolution and Function of Serine Proteases from the Mast Cell Chymase Locus
Open this publication in new window or tab >>Sculpted through Time: Evolution and Function of Serine Proteases from the Mast Cell Chymase Locus
2006 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Immune cells like NK cells, T cells, neutrophils and mast cells store high amounts of granule serine proteases, graspases. Graspases are encoded from the mast cell chymase locus. The human locus holds four genes: α-chymase, cathepsin G, and granzymes H and B. In contrast, the mouse locus contains at least 14 genes. Many of these belong to subfamilies not found in human, e.g. the Mcpt8-family. These differences hamper functional comparisons of graspases and of immune cells in the two species. Studies of the mast cell chymase locus are therefore important to better understand the mammalian immune system.

In this thesis, the evolution of the mast cell chymase locus was analysed by mapping the locus in all available mammalian genome sequences. It was revealed that one single ancestral gene founded this locus probably over 215 million years ago. This ancestor was duplicated more than 185 million years ago. One copy evolved into the α-chymases, whereas the second copy founded the families of granzymes B and H, cathepsin G, Mcpt8 and duodenases. Different subfamilies were later remarkably expanded in particular mammalian lineages, e.g. the Mcpt8- and Mcpt2-subfamilies in the rat. Four novel members of these families were identified in rat mucosal mast cells. Rat and mouse mast cells express numerous different graspases, whereas human and dog mast cells express only one graspase, chymase. To better understand mast cell functions in these species, one member of the mouse Mcpt8-family, mMCP-8, and human and dog chymase were studied. The preferred substrate sequence was analysed by substrate phage display. mMCP-8 remains yet enigmatic, although it is probably proteolytically active. Dog and human chymase, interestingly, have common preferences in certain substrate positions, but differ in others. These two chymases may have coevolved with an in vivo substrate that is conserved only in the positions with a common preference. We also obtained evidence that substrate positions on either side of the scissile bond influence each other. This kind of interactions can only be detected with a method investigating both sides simultaneously, such as substrate phage display.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2006. 90 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 250
Immunology, immune system, mast cell, chymase locus, serine protease, evolution, gene duplication, Immunologi
urn:nbn:se:uu:diva-7379 (URN)91-554-6748-2 (ISBN)
Public defence
2006-12-20, C4:305, BMC, Husargatan 3, Uppsala, 09:00
Available from: 2006-11-29 Created: 2006-11-29Bibliographically approved

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