Manzara T, Carrasco P, Gruissem W
Department of Plant Biology, University of California, Berkeley 94720.
Plant Cell. 1991 Dec;3(12):1305-16. doi: 10.1105/tpc.3.12.1305.
The five genes encoding ribulose-1,5-bisphosphate carboxylase (rbcS) from tomato are differentially expressed. Transcription of the genes is organ specific and developmentally regulated in fruit and light regulated in cotyledons and leaves. DNase I footprinting assays were used to map multiple sites of DNA-protein interaction in the promoter regions of all five genes and to determine whether the differential transcriptional activity of each gene correlated with developmental or organ-specific changes in DNA-protein interactions. We show organ-specific differences in DNase I protection patterns, suggesting that differential transcription of rbcS genes is controlled at least in part at the level of DNA-protein interactions. In contrast, no changes were detected in the DNase I footprint pattern generated with nuclear extracts from dark-grown cotyledons versus cotyledons exposed to light, implying that light-dependent regulation of rbcS transcription is controlled by protein-protein interactions or modification of DNA binding proteins. During development of tomato fruit, most DNA-protein interactions in the rbcS promoter regions disappear, coincident with the transcriptional inactivation of the rbcS genes. In nuclear extracts from nonphotosynthetic roots and red fruit, the only detectable DNase I protection corresponds to a G-box binding activity. Detection of other DNA binding proteins in extracts from these organs and expression of nonphotosynthetic genes exclude the possibility that roots and red fruit are transcriptionally inactive. The absence of complex promoter protection patterns in these organs suggests either that cooperative interactions between different DNA binding proteins are necessary to form functional transcription complexes or that there is developmental and organ-specific regulation of several rbcS-specific transcription factors in these organs. The DNase I-protected DNA sequences defined in this study are discussed in the context of conserved DNA sequence motifs and previously characterized binding sites.
番茄中编码1,5 - 二磷酸核酮糖羧化酶（rbcS）的五个基因存在差异表达。这些基因的转录具有器官特异性，在果实中受发育调控，在子叶和叶片中受光照调控。利用DNA酶I足迹分析来绘制所有五个基因启动子区域中DNA - 蛋白质相互作用的多个位点，并确定每个基因的差异转录活性是否与DNA - 蛋白质相互作用中的发育或器官特异性变化相关。我们展示了DNA酶I保护模式中的器官特异性差异，这表明rbcS基因的差异转录至少部分是在DNA - 蛋白质相互作用水平上受到控制的。相比之下，用黑暗生长的子叶与光照处理的子叶的核提取物产生的DNA酶I足迹模式未检测到变化，这意味着rbcS转录的光依赖性调控是由蛋白质 - 蛋白质相互作用或DNA结合蛋白的修饰所控制的。在番茄果实发育过程中，rbcS启动子区域中的大多数DNA - 蛋白质相互作用消失，这与rbcS基因的转录失活同时发生。在非光合根和红色果实的核提取物中，唯一可检测到的DNA酶I保护对应于一种G - 盒结合活性。在这些器官的提取物中检测到其他DNA结合蛋白以及非光合基因的表达排除了根和红色果实转录无活性的可能性。这些器官中缺乏复杂的启动子保护模式表明，要么不同DNA结合蛋白之间的协同相互作用对于形成功能性转录复合物是必要的，要么这些器官中存在对几种rbcS特异性转录因子的发育和器官特异性调控。本研究中定义的DNA酶I保护的DNA序列将在保守DNA序列基序和先前表征的结合位点的背景下进行讨论。
