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Cloning and characterization of a novel NK cell-specific serine protease gene and its functional 5′-flanking sequences

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Abstract

Rat natural killer cell Met-ase-1 (RNK-Met-1) is a 30 000 M r serine protease (granzyme) found in the cytolytic granules of CD3- large granular lymphocytes (LGL) with natural killer (NK) activity. To characterize the genomic sequences responsible for the CD3- LGL-restricted expression of this gene, we screened a rat genomic library with RNK-Met-1 cDNA, and obtained bacteriophage clones that contained the RNK-Met-1 gene. The RNK-Met-1 gene comprises 5 exons and spans approximately 5.2 kilobases (kb), exhibiting a similar structural organization to a class of CTL-serine proteases with protease catalytic residues encoded near the borders of exons 2, 3, and 5. The 5′-flanking region of the RNK-Met-1 gene contains a number of putative promoter and enhancer regulatory elements and shares several regions of homology with the 5′-flanking region of the mouse perforin gene. We have prepared nested deletions from approximately 3.3 kb of the 5′-flanking region of the RNK-Met-1 gene, and inserted these upstream of the chloramphenicol acetyltransferase (CAT) reporter gene. These 5′-flanking RNK-Met-1-CAT constructs were transiently transfected into rat LGL leukemia, T-lymphoma, and basophilic leukemia cell lines.

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  1. Mark J. Smyth

    Present address: Cellular Cytotoxicity Laboratory, Austin Research Institute, Austin Hospital, Heidelberg, 3084, Victoria, Australia

Authors and Affiliations

  1. Cellular Cytotoxicity Laboratory, Austin Research Institute, Austin Hospital, Heidelberg, 3084, Victoria, Australia

    Mark J. Smyth, Mark D. Hulett, Kevin Y. T. Thia & Joseph A. Trapani

  2. Laboratory of Experimental Immunology, Biological Response Modifiers Program, NCI-Frederick Cancer Research and Development Center, 21702-1201, Frederick, MD, USA

    Howard A. Young

  3. Biological Carcinogenesis and Development Program, PRI/DynCorp, NCI-Frederick Cancer Research and Development Center, 21702-1201, Frederick, MD, USA

    Thomas J. Sayers & Clive R. D. Carter

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  1. Mark J. Smyth
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The nucleotide sequence data reported in this Papershave been submitted to the EMBL/GenBank nucleotide sequence database and have been assigned the accession number L38482.

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Smyth, M.J., Hulett, M.D., Thia, K.Y.T. et al. Cloning and characterization of a novel NK cell-specific serine protease gene and its functional 5′-flanking sequences. Immunogenetics 42, 101–111 (1995). https://doi.org/10.1007/BF00178584

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