探討陰道滴蟲Myb2蛋白質中調控入核運輸之區域

Abstract

Myb2調控陰道滴蟲ap65-1基因的轉錄活性，通常存在於細胞核中。本研究之目的在於尋找Myb2入核控制序列。利用免疫螢光染色方法，藉蛋白質刪短或突變技術，發現Myb2入核控制之充分序列位於區域48-143，其中以酸性胺基酸聚集區域55EEDE58為入核控制所必需，而51KF52、139NRW141與143T/S145亦可能參與Myb2之入核控制。Myb2核酸結合區域 (DNA-binding domain) R2R3中，若刪除H2、H3、H4、H5與RC4等任一結構元素，所表現之蛋白質均滯留於細胞質中；顯示分子結構之完整性可能亦為Myb2入核所需。由西方墨點法偵知，144-179區域可能為轉譯後修飾之標的；此修飾可能發生於細胞質中，與Myb2之入核無關。藉由有限的激酶抑制劑處理，無法偵測得Myb2入核的訊息傳遞路徑，而Myb2入核亦不受到因生長所引起之培養環境變化的影響。這些結果顯示陰道滴蟲Myb2的入核信號 (Nuclear Localization Signal)複雜，並非由單一序列所控制。

Myb2, which regulates iron inducible transcription of the ap65-1 gene in the protozoan parasite Trichomonas vaginalis, was persistently detected in the nucleus. In the study, the nuclear localization signal of Myb2 was investgated exploiting immuneflouorescence assay to monitor subcellular localization of Myb2 and its mutant proteins. The region 48-143 was found to be sufficient, while 55EEDE58 as well as anyone of the structure elements in the R2R3 DNA-binding domain to be essential for nuclear import. Moreover, 51KF52, 139NRW141, and 143T/S145 together may also play important roles in Myb2 nuclear import. As examined by Western blotting, the region spanning 144-179 may contain a site for post-translational modification, which is not required for Myb2 nuclear import. Signal transduction pathway involve in Myb2 nuclear import were not identified. These observations suggest that Myb2 nuclear translocation is controlled by a novel nuclear localization signal.