| Project/Area Number |
63480125
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
General medical chemistry
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| Research Institution | Osaka University |
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Principal Investigator |
TANAKA Takehiko Osaka Univ. Faculty of Med. Prof., 医学部, 教授 (60028272)
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| Co-Investigator(Kenkyū-buntansha) |
NOGUCHI Tamio Osaka Univ. Faculty of Med. Assist. Prof., 医学部, 講師 (70135721)
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| Project Period (FY) |
1988 – 1989
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥6,100,000 (Direct Cost: ¥6,100,000)
Fiscal Year 1989: ¥2,700,000 (Direct Cost: ¥2,700,000)
Fiscal Year 1988: ¥3,400,000 (Direct Cost: ¥3,400,000)
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| Keywords | Pyruvate kinase isozymes / Tissue-specific expression / Cis-acting element / CAT assay / Enhancer / Hepatocyte-specific transcription factor / Insulin / L型ピルビン酸キナ-ゼ / ピルビン酸キナーゼアイソザイム / シス作用制御領域 |
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| Research Abstract |
Cis-acting elements responsible for expression of pyruvate kinase L and M genes were studied by transient DNA transfer experiments with chloramphenicol acetyltransferase (CAT) fusion genes. We found two positive regulatory regions required for expression of the L-type isozyme in adult rat hepatocytes. These regions, designated as PKL-I and PKL-II, were located between nucleotides -94 and -76, and -170 and -126, respectively. PKL-II enhanced expression from heterologous as well as homologous promoters in a manner that was independent of its orientation and position relative to the cap site. PKL-I also enhanced the promoter activity. PKL-I and PKL-II showed synergistic enhancer activity. PKL-II contains two notable sequences; one is the sequences that is reported to be the LF-AL binding site and the other is palindrome sequences. PKL-I consists of palindrome sequences which contain a sequence homologous to the LF-Bl binding site. The activities of PKL-I and PKL-II were not detected in K562 erythroleukemia cells, suggesting that PKL-I and PKL-II are very important enhancer elements for hepatocytespecific expression of the L-type isozyme.
It was also suggested that the 5'-flanking region of the Retype cap site of the L gene contained cis-acting element responsible for expression of the R-type. The 5'-flanking region of the M gene was suggested to contain two cis-acting elements.
Studies on transgenic mice containing CAT fusion genes indicated that the 5'-flanking region of the L-type cap site up to -3000 contained element responsive to insulin.
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