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Expansion of the mast cell chymase locus over the past 200 million years of mammalian evolution
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.
2006 (English)In: Immunogenetics, ISSN 0093-7711, E-ISSN 1432-1211, Vol. 58, no 8, 655-669 p.Article in journal (Refereed) Published
Abstract [en]

The acidic granules of natural killer (NK) cells, T cells, mast cells, and neutrophils store large amounts of serine proteases. Functionally, these proteases are involved, e.g., in the induction of apoptosis, the recruitment of inflammatory cells, and the remodeling of extra-cellular matrix. Among the granule proteases are the phylogenetically related mast cell chymases, neutrophil cathepsin G, and T-cell granzymes (Gzm B to H and Gzm N), which share the characteristic absence of a Cys191–Cys220 bridge. The genes of these proteases are clustered in one locus, the mast cell chymase locus, in all previously investigated mammals. In this paper, we present a detailed analysis of the chymase locus in cattle (Bos taurus) and opossum (Monodelphis domestica). The gained information delineates the evolution of the chymase locus over more than 200 million years. Surprisingly, the cattle chymase locus contains two α-chymase and two cathepsin G genes where all other studied chymase loci have single genes. Moreover, the cattle locus holds at least four genes for duodenases, which are not found in other chymase loci. Interestingly, duodenases seem to have digestive rather than immune functions. In opossum, on the other hand, only two chymase locus-related genes have been identified. These two genes are not arranged in one locus, but appear to have been separated by a marsupial-specific chromosomal rearrangement. Phylogenetic analyses place one of the opossum genes firmly with mast cell α-chymases, which indicates that the α-chymase had already evolved as a separate, clearly identifiable gene before the separation of marsupials and placental mammals. In contrast, the second gene in opossum is positioned phylogenetically between granzymes, cathepsin G, and the duodenases. These genes, therefore, probably evolved as separate subfamilies after the separation of placental mammals from marsupials. In platypus, only one chymase locus-like sequence could be identified. This previously published “granzyme” does not cluster clearly with any of the chymase locus gene families, but shares the absence of the Cys191–Cys220 bridge with the other chymase locus proteases. These findings indicate that all chymase locus genes are derived from a single ancestor that was present more than 200 million years ago.

Place, publisher, year, edition, pages
2006. Vol. 58, no 8, 655-669 p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-95243DOI: 10.1007/s00251-006-0126-1OAI: oai:DiVA.org:uu-95243DiVA: diva2:169381
Available from: 2006-11-29 Created: 2006-11-29 Last updated: 2017-12-14Bibliographically 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.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 250
Keyword
Immunology, immune system, mast cell, chymase locus, serine protease, evolution, gene duplication, Immunologi
Identifiers
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
Opponent
Supervisors
Available from: 2006-11-29 Created: 2006-11-29Bibliographically approved
2. Hematopoietic Serine Proteases from the Mast Cell Chymase and Tryptase Loci - a Functional and Evolutionary Analysis
Open this publication in new window or tab >>Hematopoietic Serine Proteases from the Mast Cell Chymase and Tryptase Loci - a Functional and Evolutionary Analysis
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Mast cells are key effector cells in allergic and inflammatory diseases. However, their primary role is most likely in host defence against parasitic and bacterial infections. Mast cells are a particularly rich source of serine proteases. These proteases belong to the chymase or the tryptase family, which are encoded from the mast cell chymase and the multigene tryptase loci, respectively. To better understand the biological functions and the molecular evolution of these enzymes we have studied the organisation of these two loci in species ranging from fish to human. We show that the mast cell chymase locus has evolved from a single founder gene to a complex locus during the past 200 Myr of mammalian evolution. Forty-five fish candidate genes for hematopoietic serine proteases were also identified. However, in phylogenetic analyses none of them grouped with individual branches holding mammalian mast cell chymase locus genes, indicating an independent parallel evolution in fish.

Studies of the evolution of the multigene tryptase locus showed that this locus has been highly conserved between marsupials and eutherians. However, no genes belonging to the individual subfamilies identified in eutherians could be identified in fish, amphibians or in birds, which also here indicates parallel evolution.

To study the evolution of specific cleavage specificities associated with these proteases, the extended cleavage specificity of opossum α-chymase was determined and found to be nearly identical to human mast cell chymase and the major mouse mast cell chymase mMCP-4. This indicates a strong pressure to maintain this specificity during mammalian evolution.

Basophils are rare blood cells with functions similar to mast cells that when mature almost completely lack mRNA. To study the proteome and to primarily characterize the granule protein content of basophils, an in vitro purification protocol was developed to obtain transcriptionally active umbilical cord blood-derived basophil precursors.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2008. 72 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 427
Keyword
Cell and molecular biology, immune system, mast cell, basophil, mast cell chymase locus, multigene tryptase locus, serine protease, evolution, Cell- och molekylärbiologi
Identifiers
urn:nbn:se:uu:diva-8676 (URN)978-91-554-7179-8 (ISBN)
Public defence
2008-05-16, C10:305, BMC, Husargatan 3, Box 596, 75124 Uppsala, 09:15
Opponent
Supervisors
Available from: 2008-04-25 Created: 2008-04-25 Last updated: 2011-07-08Bibliographically approved

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Gallwitz, MaikeReimer, JennyHellman, Lars

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