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Title :M12-Tet-Off pTRE2pur FLAG mTRAF3細胞株の樹立
Authors :田邊, 香野
坂本, 瞳
乾, 誠治
Issue Date :30-Mar-2013
Citation jtitle :熊本大学医学部保健学科紀要
vol. :9
start page :27
end page :38
Abstract :IL-4 induces IgE class switching in B cells together with CD40 stimulation. Studies on the signal transduction mechanism downstream of IL-4 receptor have revealed that NF-κB as well as Jak-STAT pathway plays an important role. We showed the possibility that alternative pathway of NF-κB was activated after IL-4 stimulation of murine B cell line M12. Alternative pathway of NF-κB is strictly regulated by constant degradation of NIK, an essential molecule for NF-κB activation. TRAF3 is the key molecule to keep the alternative pathway in an inactive state. TRAF3 is also regulated by the constant degradation by ubiquitin-proteasome system. To elucidate the mechanism of the TRAF3 degradation, we prepared Tet-Off gene expression system in M12 to inducibly express TRAF3 which was toxic when overexpressed in cells. Firstly, pTet-Off vector was introduced into M12 cells by electroporation and G418 resistant clones were selected which harbored pTet-Off. Clones were further selected to show high responsiveness to Doxycyclin withdrawal using transient expression of luciferase vector. One of the clones, C'' , was transfected with pTRE2pur-mTRAF3 vector to obtain FLAG-tagged mTRAF3 expressing cells upon induction. One clone, C'' -1' , was successfully established which expressed mTRAF3 in an inducible manner.
Type Local :紀要論文
ISSN :18807151
Publisher :熊本大学
URI :http://hdl.handle.net/2298/27461
Appears in Collections:Bulletin of Kumamoto University, School of Health Sciences (Medicine)
Please use this identifier to cite or link to this item: http://hdl.handle.net/2298/27461