| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1999: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1998: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Research Abstract |
Peptidylarginine deiminases (PAD), a group of post-translational enzymes, catalyze the conversion of protein-bound arginine residues to citrulline residues in a calcium ion dependent manner and are widely distributed in various organs of vertebrates. We found the existence of four isoforms of PAD (types I, II, III, and IV) in mouse. However, the relative functional consequences of the isoforms with respect to their co-location in tissues have yet to be explored. Recently we succeeded to clone the full-length cDNAs of every mouse PAD isoforms and deduced their primary structures. In order to elucidate the functions of each PAD isoform in murine cells, we make strategies by using knockout mouse whose PAD gene(s) is (are) destroyed by replacing the PAD gene to a vector harboring a partial of the PAD gene and a resistant gene against an antibiotic. From 1998 to 1999 we succeeded to clone the genes of every PAD isoform and determined their nucleotide sequences of 5'-flanking region, and we also extended the cloning of long region of each PAD isoform from bacterial artificial chromosome (BAC) libraries of embryonic stem cell lines (ES-129/SvJ). In this year, the found clone was derived from the type II gene and was useful to make targeting vectors for homologous recombination of PAD genes. Therefore, we focused on the PAD types II gene for the knockout experiment and succeeded to isolate the recombinant ES cells. Now, we are establishing the knock out mouse whose PAD type II gene is destroyed.
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