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Title: Cloning and characterization of a novel NK cell-specific serine protease gene and its functional 5'-flanking sequences.
Austin Authors: Smyth, Mark J;Hulett, M D;Thia, K Y;Young, H A;Sayers, T J;Carter, C R;Trapani, Joseph A
Affiliation: Cellular Cytotoxicity Laboratory, Austin Research Institute, Austin Hospital, Heidelberg, Victoria, Australia
Issue Date: 16-May-1995
Publication information: Immunogenetics; 42(2): 101-11
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. The transcriptional activity of the RNK-Met-1 5'-flanking region was strong, restricted to the RNK-16 LGL leukemia and controlled by several positive cis-acting regions spread over at least 3.3 kb. The longest and most active 5'-flanking region (-3341 to -33) was also used to drive specific expression of beta-galactosidase in RNK-16. These data are consistent with the NK cell-specific expression of RNK-Met-1 and suggest the potential utility of this gene promoter in the development of transgene models of NK cell biology in vivo.
Gov't Doc #: 7607701
Type: Journal Article
Subjects: Amino Acid Sequence
Base Sequence
Blotting, Southern
Cloning, Molecular
Killer Cells, Natural.enzymology
Molecular Sequence Data
Serine Endopeptidases.chemistry.genetics
Appears in Collections:Journal articles

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