Natural killer (NK) cells, lymphocytes of the innate immune system, can recognize a wide array of cells via several receptors families such as Ly49 and NKR-P1. The Nkrp1 gene family encode for C-type lectin-like receptors which can recognize their ligands, Clr, on target cells. Nkrp1 and Clr genes are intertwined in the NK gene complex and are thus inherited together. The Nkrp1-Clr genes in 129S6 and BALB/c mouse strains show significant sequence polymorphism compared to those of C57BL/6 mice while the overall gene organization and gene number are conserved. RT-PCR was utilized to study the expression of individual Nkrp1-Clr genes. In situ hybridization was performed to validate expression results from RT-PCR, as well as to verify the cell types in which Nkrp1-Clr genes are expressed. Surprisingly, our expression studies reveal an interesting pattern of expression of Nkrp1 and Clr genes not only in lymphoid tissues but also in the epithelial cells of the intestine, kidney, eye and lung, the myocytes of the heart and skeletal muscle, and possibly some endothelial cells, indicating novel functions of NK cells in these tissues.

Natural killer (NK) cells, lymphocytes of the innate immune system, can recognize a wide array of cells via several receptors families such as Ly49 and NKR-P1. The Nkrp1 gene family encode for C-type lectin-like receptors which can recognize their ligands, Clr, on target cells. Nkrp1 and Clr genes are intertwined in the NK gene complex and are thus inherited together. The Nkrp1-Clr genes in 129S6 and BALB/c mouse strains show significant sequence polymorphism compared to those of C57BL/6 mice while the overall gene organization and gene number are conserved. RT-PCR was utilized to study the expression of individual Nkrp1-Clr genes. In situ hybridization was performed to validate expression results from RT-PCR, as well as to verify the cell types in which Nkrp1-Clr genes are expressed. Surprisingly, our expression studies reveal an interesting pattern of expression of Nkrp1 and Clr genes not only in lymphoid tissues but also in the epithelial cells of the intestine, kidney, eye and lung, the myocytes of the heart and skeletal muscle, and possibly some endothelial cells, indicating novel functions of NK cells in these tissues.

Natural killer (NK) cells, lymphocytes of the innate immune system, can recognize a wide array of cells via several receptors families such as Ly49 and NKR-P1. The Nkrp1 gene family encode for C-type lectin-like receptors which can recognize their ligands, Clr, on target cells. Nkrp1 and Clr genes are intertwined in the NK gene complex and are thus inherited together. The Nkrp1-Clr genes in 129S6 and BALB/c mouse strains show significant sequence polymorphism compared to those of C57BL/6 mice while the overall gene organization and gene number are conserved. RT-PCR was utilized to study the expression of individual Nkrp1-Clr genes. In situ hybridization was performed to validate expression results from RT-PCR, as well as to verify the cell types in which Nkrp1-Clr genes are expressed. Surprisingly, our expression studies reveal an interesting pattern of expression of Nkrp1 and Clr genes not only in lymphoid tissues but also in the epithelial cells of the intestine, kidney, eye and lung, the myocytes of the heart and skeletal muscle, and possibly some endothelial cells, indicating novel functions of NK cells in these tissues.

Natural killer cells play a significant role in the immune response against tumor and infected cells. NK cells express a wide variety of surface receptors, including NKRP1, a C-type lectin-like family of both activating and inhibitory receptors. Recently, ligands have been found for some of these previously orphan molecules, some of them lying within the same family. This is also the case of rat Clr-b as a cognitive ligand for rat NKRP1B. It has been shown that in rat, this inhibitory NKRP1B-Clr-b mutual receptor system is subverted by rat cytomegalovirus protein RCTL, a viral version of Clr-b, which serves as a decoy ligand for NK cells. The aim of my diploma thesis was cloning and production of the above mentioned C-type lectin-like proteins based on transient transfection of HEK293T cell line in a suspension culture. This expression system allows us not only to obtain proteins of our interest with a satisfactory yield but also in their native conformation, removing the need for time consuming and often fruitless refolding procedures required in case of using the E. coli expression system. Success was achieved in case of Clr-b and NKRP1B receptors from both WAG and SD strains. Proteins were purified using IMAC followed by gel filtration, identified by mass spectrometry and characterized by disulfide...