O'Donnell K H, Chen C T, Wensink P C
Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254-9110.
Mol Cell Biol. 1994 Sep;14(9):6398-408. doi: 10.1128/mcb.14.9.6398-6408.1994.
We identify DNA regions that are necessary for the ubiquitous expression of the Drosophila melanogaster alpha 1-tubulin (alpha 1t) gene. In vitro transcription showed that two upstream regions, tubulin element 1 (TE1 [29 bp]) and tubulin element 2 (TE2 [68 bp]), and a downstream region activate transcription. Germ line transformation demonstrated that these three regions are sufficient to direct the alpha 1t core promoter to begin transcribing at the stage of cellular blastoderm formation and to continue thereafter at high levels in all tissues and developmental stages. Remarkably, mutation of any one of these regions results in high sensitivity to chromosomal position effects, producing different but reproducible tissue-specific patterns of expression in each transformed line. None of these regions behaves as an enhancer in a conventional germ line transformation test. These observations show that these three regions, two of which bind the GAGA transcription factor, act ubiquitously to insulate from position effects and to activate transcription. The results also provide vectors for ubiquitous expression of gene products and for examining silencer activities.
我们鉴定出了黑腹果蝇α1 - 微管蛋白（α1t）基因普遍表达所必需的DNA区域。体外转录显示，两个上游区域，即微管蛋白元件1（TE1 [29 bp]）和微管蛋白元件2（TE2 [68 bp]），以及一个下游区域可激活转录。生殖系转化表明，这三个区域足以指导α1t核心启动子在细胞胚盘形成阶段开始转录，并在随后的所有组织和发育阶段持续高水平转录。值得注意的是，这些区域中的任何一个发生突变都会导致对染色体位置效应高度敏感，在每个转化系中产生不同但可重复的组织特异性表达模式。在传统的生殖系转化试验中，这些区域均未表现出增强子的作用。这些观察结果表明，这三个区域中的两个与GAGA转录因子结合，它们普遍发挥作用，以隔离位置效应并激活转录。这些结果还提供了用于基因产物普遍表达和检查沉默子活性的载体。
