Identification of the transcription factors binding to the negative element of K14 promoter in TPA-treated C9 Cell

• Main Authors: Wu, Shao-Wen, 吳少文
• Other Authors: Chao, Chung-Faye
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/73712466232258167049

碩士 === 國防醫學院 === 生物及解剖學研究所 === 92 === Keratin 14 (K14) is expressed in the basal cell layer of epidermis, and then down-regulative expressed when cell migrate toward the upper layer. This down regulation phenomenon of K14 is necessary for the epidermal cell differentiation hence K14 is considered as an epidermal cell differentiation marker. TPA can induce down regulation expression of K14 in cultured keratinocytes and epidermal carcinoma cell lines. Through using deletion mutant of K14 promoter fragment and gel shift assay, it was demonstrated that TPA response and non-response elements of K14 promoter were located at the region of -379~ -344 and -405~ -380 resparately. For identification of the different transcription factors, which bind to the negative element of K14 promoter in C9 cell with/without treatment of TPA, extracted nuclear proteins were incubated with magnetic beads ligated with designed DNA probes. Then the DNA binding proteins were eluted and analyzed by SDS-PAGE. The different bands were cut out of gel and analyzed by mass spectrometer. In this study, we found that the proteins binding to the negative element of K14 promoter at the region of -405~-344 of TPA-treated C9 cell are more than non-TPA treated C9 cell. In TPA-treated C9 cell, β-actin may involve in the formation of those nuclear binding proteins complex at this region. Other unknown novel proteins may also contribute to the regulation of K14 promoter and need further investigation.