Serra E, Zemzoumi K, Trolet J, Capron A, Dissous C
Unité INSERM 167, Institut Pasteur, Lille, France.
Mol Biochem Parasitol. 1996 Dec 2;83(1):69-80. doi: 10.1016/s0166-6851(96)02751-x.
The ability of the 5' flanking region of the gene encoding the 28 kDa glutathione S-transferase of Schistosoma mansoni gene to promote transcription, was studied in different mammalian cell lines. Results of transient transfection assays showed a strong activity of the -277 to +1 nt region of the Sm28GST gene, comparable to that of well-studied promoters. Deletion analysis indicated that an AP-1 site and two closely located CCAAT (Y1 and Y2) boxes were the principal motifs responsible for the promoter activity. Binding of the NF-Y complex to Y1 and Y2, as well as to a third CCAAT box (Y3) close to the promoter TATA box, was compared in gel shift and super-shift experiments. All of the three Y boxes bound protein complexes from S. mansoni nuclear extracts that were shown to contain the A subunit of the schistosome NF-Y complex (SMNF-YA). Competition assays revealed a differential affinity of the Y1, Y2 and Y3 sequences for NF-Y. The Y1, Y2 and Y3 regions were also shown to activate transcription when included in an heterologous promoter and data obtained strongly suggested the involvement of SMNF-Y in multimeric complexes during this process.
研究了曼氏血吸虫28 kDa谷胱甘肽S-转移酶基因编码区5'侧翼区域在不同哺乳动物细胞系中促进转录的能力。瞬时转染试验结果显示，Sm28GST基因-277至+1 nt区域具有很强的活性，与研究充分的启动子相当。缺失分析表明，一个AP-1位点和两个紧密相邻的CCAAT（Y1和Y2）框是负责启动子活性的主要基序。在凝胶迁移和超迁移实验中，比较了NF-Y复合物与Y1和Y2以及启动子TATA框附近的第三个CCAAT框（Y3）的结合情况。所有这三个Y框都能结合来自曼氏血吸虫核提取物的蛋白质复合物，这些复合物被证明含有血吸虫NF-Y复合物（SMNF-YA）的A亚基。竞争试验揭示了Y1、Y2和Y3序列对NF-Y的不同亲和力。当Y1、Y2和Y3区域包含在异源启动子中时，也显示出能激活转录，并且所获得的数据有力地表明在此过程中SMNF-Y参与了多聚体复合物的形成。
