Abstract
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.