金屬感應轉錄因子為鋅指結構之轉錄因子，它能與金屬硫蛋白啟動子上的金屬感應序列結合，藉以調控金屬硫蛋白基因的表現。當細胞受到金屬鋅的刺激後，金屬感應轉錄因子會快速地由細胞質進入細胞核中，並且活化蛋白質的轉錄作用。由實驗室先前的研究發現金屬感應因子在其C端具有能被SUMO修飾的IKQE保留序列，為了研究金屬感應轉錄因子被SUMO修飾後的特性，我們將其DNA結合區以Gal4 DNA結合區置換，形成Gal4-TAD融合蛋白質，此融合蛋白質主要分布於細胞核中。在本篇研究中發現Gal4-TAD能夠被SUMO修飾，而當突變SUMO保留序列後，會增加蛋白質的轉錄活性，表示SUMO化會抑制蛋白質的轉錄活性，也顯示SUMO化的發生和金屬感應轉錄因子的完整結構無關，而是藉由辨識SUMO保留序列進行修飾。為了研究金屬感應轉錄因子被SUMO修飾的程度是否和其在細胞中的分布有關，因此我們突變Gal4-TAD蛋白質上的核定位信號使其分布於細胞質中。由實驗結果發現無論蛋白質是位於細胞核或是細胞質皆能被SUMO修飾，而且其修飾的程度相似。當細胞以鋅處理後，在細胞核或是細胞質中皆能觀察到蛋白質被SUMO修飾的程度隨著時間與劑量而有逐漸遞減的現象，表示金屬能夠調控金屬感應轉錄因子被SUMO修飾的程度。當細胞以鋅處理後，SENP1和SENP2蛋白質的量並未隨之改變，同時RanGAP1被SUMO修飾的程度也未受影響，表示金屬造成金屬感應轉錄因子之去SUMO化作用是具有專一性的。

Metal responsive transcription factor 1 (MTF-1) is a zinc finger protein that recognizes metal responsive elements in the promoter region. MTF-1 retains mainly in the cytoplasm. Upon stimulation, MTF-1 translocates from the cytoplasm into the nucleus and initiates the transcription. Recently, we have demonstrated the occurrence of SUMOylation in MTF-1 and the site of modification was identified. To characterize the SUMOylation in MTF-1, the zinc finger domain of MTF-1 was replaced by Gal4 DNA binding domain (Gal4-TAD). This fusion protein locates in the nucleus when synthesized. The transcriptional activity was enhanced significantly once the site for SUMOylation was mutated, indicating the repression characteristics of SUMOylation for this transcription factor. The result also shows the independence of SUMOylation with zinc finger motifs. To explore whether the SUMOylation is related to the localization of MTF-1, the nuclear localization signal of Gal4-TAD was mutated such that the protein retained only in the cytoplasm. Proteins in either the cytoplasm or the nucleus can be SUMOylated, indicating the irrelevance of cellular location for the modification. When treating cells with zinc, the level of SUMOylation declined in either cytoplasm or nucleus. This finding indicates that metal treatment, but not the translocation process, is the critical factor to remove SUMO from MTF-1. Addition of metal apparently did not affect the cellular level of SUMO-specific protease, SENP1 and SENP2. The metal-induced reduction in SUMO modification did not occur in RanGAP1. Other mechanism should involve in attenuating the SUMOylation of MTF-1 by metals. These results indicate that the reduction of MTF-1 SUMOylation is not a nonspecific response by metal treatment.

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