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GENOMIC MAPPING AND PROMOTER CHARACTERIZATION OF THE OVINE TROPHOBLAST KUNITZ DOMAIN PROTEIN-1 (ovTKDP-1).
Chakrabarty, Anindita1, Green, Jonathan2, MacLean, James3, Roberts, Robert1,2,3, 1 2 3
ABSTRACT- The trophoblast Kunitz domain proteins (TKDP) are secretory proteins produced by trophoblast mononuclear cells of cattle and sheep. Each of these proteins contains a classical Kunitz-type serine protease inhibitory domain at the C-terminus and one or more N-terminal domains of approximately 80 amino acids. Most of the TKDP are highly expressed in tightly regulated, narrow windows during pregnancy. The cDNA for ovTKDP-1 encodes a protein with two N-terminal domains (D1 and D2) preceding the Kunitz domain. The ovTKDP-1 gene was obtained by stringent hybridization screening of a lambda EMBL-3 genomic library with a probe for D1 and D2. Mapping of the gene was performed by PCR with different combinations of cDNA-specific primers, followed by sequencing. The Kunitz domain and the signal peptide/propeptide are encoded by single exons. Each of the N-terminal domains is encoded by three short exons. Curiously, two of the exons in D2 and all three exons in D1 have many features typical of eukaryotic introns, suggesting that these exons evolved recently from introns. Parts of the intron sequences between the exons comprising D1 and D2 also exhibit high sequence identity, indicating that the two repeat domains of ovTKDP-1 might have arisen from a recent duplication. Interferon tau is important for maternal recognition of pregnancy in ruminant ungulates. Interestingly, ovTKDP-1 and interferon-tau exhibit exactly the same expression pattern and tissue specificity. For this reason, we sought to compare the transcriptional regulation of these two proteins. The 5′UTR of the ovTKDP-1 gene has been sequenced and mapped to identify putative transcription factor binding sites. However, no obvious similarities with the ovine interferon tau gene were observed, suggesting that the transcriptional controls operating on the two genes differ. Supported by NIH grant HD35898.
KEY WORDS: trophoblast Kunitz domain proteins, introns, exons, interferon-tau
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