MTF-1是一個具有六個C2H2類型zinc finger的轉錄蛋白，能與MT啟動子上的MRE序列 (TGCRCNC) 結合，藉此調控MT基因的基礎表現，或是金屬誘導時的表現。已知在細胞受到鋅刺激後，MTF-1會由快速地由細胞質進入細胞核，然而，MTF-1以何作為核定位信號，至今仍是不清楚的。在我們的實驗中，將MTF-1的片段與eGFP融合，利用螢光顯微鏡與西方墨點法分析其在細胞中的分佈。結果發現將MTF-1的前兩個zinc finger與後四個zinc finger分別與eGFP融合，其融合蛋白都會累積於細胞核。因此推測MTF-1的zinc finger區域中可能具有兩個功能獨立的NLS。此外，我們也更進一步地證實它們的核定位作用是不需要zinc finger三級結構的完整性以及與DNA的結合活性。我們也觀察到MTF-1的 N端序列 (1-136 amino acid) 可能會抑制核定位作用。我們另外再利用點突變的方法破壞zinc finger的結構，以觀察各個zinc finger在金屬誘導MTF-1進核的過程中的重要性，同時也發現zinc finger的三級結構可能參與MTF-1進核的調控。綜合上述的實驗的結果，指出MTF-1利用zinc finger中的兩個NLS進入細胞核，並藉由N端序列以及六個zinc finger的共同調控MTF-1的進核。

Metal-responsive transcription factor 1 (MTF-1) is a transcription factor that contains six C2H2¬¬ zinc fingers and is essential for basal and heavy metal-induced MT gene expression by binding to MRE (TGCRCNC) in the MT promoter. It was previously shown that MTF-1 rapidly translocates to the nucleus in response to zinc exposure. However, regions within the MTF-1 protein that serve as nuclear localization signal (NLS) remain unclear. In our study, enhanced green fluorescent protein (eGFP) was fused with MTF-1, and their subcellular localization was examined by fluorescence microscopy and Western blot. Fusion of eGFP with the first two zinc fingers of MTF-1 and with the last four zinc fingers of MTF-1 respectively led the protein to accumulate in the nucleus. Thus, we suggest that there may be two NLS with independent functions in MTF-1. Furthermore, we demonstrated neither the tertiary structure of the zinc fingers nor the DNA binding activity of the protein are necessary for nuclear localization. We also noted that the N-terminal domain of MTF-1 (1-136 amino acid) appears to inhibit nuclear localization. We create the mutant of MTF-1 containing histidine to alanine mutations that disrupt the structure of each zinc individually to identify the different significance of each zinc figure. At the same time, we observe tertiary structure of zinc finger also involved in regulating nuclear import. These data suggest that two nuclear localization signals in the zinc finger domain direct MTF-1 to nucleus and that N-terminal domain and six zinc fingers regulate nuclear import of MTF-1 cooperatively.

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