Abstract
We investigated the NF-κB-like factor induced in the late-passage human oral mucosal fibroblasts stimulated with interleukin-1 (IL-1). Compared with the NFκBs of HeLa cells and early-passage fibroblasts, the NF-κB-like factor of late-passage (passage 15) fibroblasts migrated faster in the electrophoretic mobility shift assay (EMSA) and behaved like a 70-80 kDa protein in the gel filtration chromatography. Both antibodies against p50 and p65 subunits of NF-κB could supershift the small NF-κB-like factor of late- passage cells in the EMSAs. A 47-kDa band was detected in late-passage fibroblasts by immunoblotting against p50. The mobility of the trypsin- degraded NF-κB of HeLa cells corresponded to that of the small NF-κB-like factor of late-passage fibroblasts in the EMSAs. Furthermore, when the nuclear extracts of the IL-1-stimulated HeLa cells were incubated with those of the IL-1-stimulated old fibroblasts, the p65-p50 NF-κB band disappeared, leaving behind a small NF-κB-like band. This reduction of NF-κB was prevented by the addition of a cysteine protease inhibitor leupeptin. These results suggest that the small NF-κB-like factor of late-passage fibroblasts is a part of the NF-κB truncated by aging-induced protease(s).
| Original language | English |
|---|---|
| Pages (from-to) | 247-255 |
| Number of pages | 9 |
| Journal | Journal of cellular physiology |
| Volume | 182 |
| Issue number | 2 |
| DOIs | |
| Publication status | Published - Jan 13 2000 |
| Externally published | Yes |
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All Science Journal Classification (ASJC) codes
- Physiology
- Clinical Biochemistry
- Cell Biology
Cite this
Aging-dependent proteolysis of NF-κB in human fibroblasts. / Ikebe, Tetsuro; Jimi, Eijiro; Beppu, Mahiro; Takeuchi, Hiroshi; Nakayama, Hideki; Shirasuna, Kanemitsu.
In: Journal of cellular physiology, Vol. 182, No. 2, 13.01.2000, p. 247-255.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Aging-dependent proteolysis of NF-κB in human fibroblasts
AU - Ikebe, Tetsuro
AU - Jimi, Eijiro
AU - Beppu, Mahiro
AU - Takeuchi, Hiroshi
AU - Nakayama, Hideki
AU - Shirasuna, Kanemitsu
PY - 2000/1/13
Y1 - 2000/1/13
N2 - We investigated the NF-κB-like factor induced in the late-passage human oral mucosal fibroblasts stimulated with interleukin-1 (IL-1). Compared with the NFκBs of HeLa cells and early-passage fibroblasts, the NF-κB-like factor of late-passage (passage 15) fibroblasts migrated faster in the electrophoretic mobility shift assay (EMSA) and behaved like a 70-80 kDa protein in the gel filtration chromatography. Both antibodies against p50 and p65 subunits of NF-κB could supershift the small NF-κB-like factor of late- passage cells in the EMSAs. A 47-kDa band was detected in late-passage fibroblasts by immunoblotting against p50. The mobility of the trypsin- degraded NF-κB of HeLa cells corresponded to that of the small NF-κB-like factor of late-passage fibroblasts in the EMSAs. Furthermore, when the nuclear extracts of the IL-1-stimulated HeLa cells were incubated with those of the IL-1-stimulated old fibroblasts, the p65-p50 NF-κB band disappeared, leaving behind a small NF-κB-like band. This reduction of NF-κB was prevented by the addition of a cysteine protease inhibitor leupeptin. These results suggest that the small NF-κB-like factor of late-passage fibroblasts is a part of the NF-κB truncated by aging-induced protease(s).
AB - We investigated the NF-κB-like factor induced in the late-passage human oral mucosal fibroblasts stimulated with interleukin-1 (IL-1). Compared with the NFκBs of HeLa cells and early-passage fibroblasts, the NF-κB-like factor of late-passage (passage 15) fibroblasts migrated faster in the electrophoretic mobility shift assay (EMSA) and behaved like a 70-80 kDa protein in the gel filtration chromatography. Both antibodies against p50 and p65 subunits of NF-κB could supershift the small NF-κB-like factor of late- passage cells in the EMSAs. A 47-kDa band was detected in late-passage fibroblasts by immunoblotting against p50. The mobility of the trypsin- degraded NF-κB of HeLa cells corresponded to that of the small NF-κB-like factor of late-passage fibroblasts in the EMSAs. Furthermore, when the nuclear extracts of the IL-1-stimulated HeLa cells were incubated with those of the IL-1-stimulated old fibroblasts, the p65-p50 NF-κB band disappeared, leaving behind a small NF-κB-like band. This reduction of NF-κB was prevented by the addition of a cysteine protease inhibitor leupeptin. These results suggest that the small NF-κB-like factor of late-passage fibroblasts is a part of the NF-κB truncated by aging-induced protease(s).
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UR - http://www.scopus.com/inward/citedby.url?scp=0033961237&partnerID=8YFLogxK
U2 - 10.1002/(SICI)1097-4652(200002)182:2<247::AID-JCP14>3.0.CO;2-S
DO - 10.1002/(SICI)1097-4652(200002)182:2<247::AID-JCP14>3.0.CO;2-S
M3 - Article
C2 - 10623889
AN - SCOPUS:0033961237
VL - 182
SP - 247
EP - 255
JO - Journal of Cellular Physiology
JF - Journal of Cellular Physiology
SN - 0021-9541
IS - 2
ER -