| Project/Area Number |
14571982
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Periodontal dentistry
|
|---|
| Research Institution | OKAYAMA UNIVERSITY |
|---|
Principal Investigator |
MYOKAI Fumio Okayama University, University Hospital of Medicine and Dentistry, Assistant Professor, 医学部・歯学部附属病院, 講師 (50263588)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
ARAI Hideo Okayama University, University Hospital of Medicine and Dentistry, Assistant Professor, 医学部・歯学部附属病院, 講師 (70222718)
NISHIMURA Fusanori Okayama University, Graduate School of Medicine and Dentistry, Associate Professor, 大学院・医歯学総合研究科, 助教授 (80208222)
TAKASHIBA Shogo Okayama University, Graduate School of Medicine and Dentistry, Professor, 大学院・医歯学総合研究科, 教授 (50226768)
KOHNO Takayuki Okayama University, University Hospital of Medicine and Dcntistry, Instructor, 医学部・歯学部附属病院, 助手 (80284074)
MAEDA Hiroshi Okayama University, Graduate School of Medicine and Dentistry, Instructor, 大学院・医歯学総合研究科, 助手 (00274001)
|
|---|
| Project Period (FY) |
2002 – 2003
|
| Project Status |
Completed (Fiscal Year 2003)
|
| Budget Amount *help |
¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 2003: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2002: ¥2,500,000 (Direct Cost: ¥2,500,000)
|
|---|
| Keywords | LITAF / promoter / Western blot / TNF-α / 転写制御 / 歯周病病態 |
|---|
| Research Abstract |
LPS-induced TNF-α factor (LITAF) is a recently identified novel transcription factor controlling TNF-α gene expression in human monocytic cells (Myokai et al., Proc Natl Acad Sci USA, 96: 4518-4523, 1999). To characterize LITAF promoter, we isolated a 1.2 kb fragment of human genomic DNA containing 5'-flanking sequence of the LITAF gene. Primer extension analysis revealed that a transcription start site situated 234 bases upstream from the translation start site in the LITAF gene. The promoter sequence contained the similar consensus sequences for AP-1 and NF-IL6 which were known to be responsible for signaling by LPS. For reporter gene assays in human T-cell leukemia cell line (Jurkat), a series of constructs with fragments of increasing length of the LITAF promoter were coupled. to the firefly luciferase gene. A 34-bp sequence domain located from nucleotides -76 to -43 in the promoter, in which the consensus binding site for AP-1 and NF-IL6 was missing, exhibited the highest reporter
… More
gene activity. Moreover, the domain did not include any known consensus sequences. These results suggest that the new sequence domain contributes the up-regulation of LITAF gene transcription.
In the following study, we aimed to examine the localization and kinetics of LITAF protein in THP-1 cells stimulated with LPS. By immunological staining using anti-LITAF monoclonal antibody, an accumulation of LITAF protein was detected in the nuclei of the LPS-stimulated cells whereas it was detected in the cytoplasm of static control cells. Western blot analysis revealed the kinetics of protein in both nuclei and cytoplasm of THP-1 cells stimulated with or without LPS. The nuclear LITAF protein was increased followed by 2 h stimulation, whereas it was recovered its basal level even by the stimulation between 2 and 24 h (student t test; p<0.05). No significant change in the cytoplasmic LITAF protein level was detected followed by the stimulation between 0.5 and 24 h. These results suggested that LITAF protein was transported form the cytoplasm to the nuclei by in the LPS stimulation. Some DNA binding proteins may serve the transportation of the LITAF protein, because the LITAF protein is lack of nuclear localization signal. Less
|
