Carter R S, Bhat N K, Basu A, Avadhani N G
Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia 19104.
J Biol Chem. 1992 Nov 15;267(32):23418-26.
DNA sequences required for expression of the mouse cytochrome c oxidase subunit IV (COXIV) promoter were identified by transient expression of recombinant COXIV-chloramphenicol acetyltransferase constructs in COS and NIH-3T3 cells. Activity of the COXIV promoter is shown to depend upon upstream Sp1 binding sequences and two tandemly repeated 21-base pair sequence elements each mapping to sites of mRNA initiation. Each initiation region repeat contains a binding site for an ets-related transcription factor which demonstrates specificity for the characteristic GGAA ets sequence motif and reactivity with an ets domain-directed monoclonal pan ets antibody. The two 21-base pair repeats are sufficient for transcriptional activity suggesting that the ets-related factor may be involved in both transcriptional activation and start site positioning. The ets-related protein found in COS nuclear extracts is shown to be identical or closely related to the GA-binding protein (GABP) by comparison of electrophoretic mobilities and immunological reactivities of DNA-protein complexes formed with purified recombinant expressed GABP alpha and beta subunits. Sp1 and the GABP-related factors also bind to another mouse cytochrome oxidase subunit gene COXVb. The similar promoter features of these two genes suggests a possible means of coordinate transcriptional regulation among such respiratory proteins.
通过在COS和NIH - 3T3细胞中瞬时表达重组COXIV - 氯霉素乙酰转移酶构建体,鉴定了小鼠细胞色素c氧化酶亚基IV(COXIV)启动子表达所需的DNA序列。结果表明,COXIV启动子的活性取决于上游Sp1结合序列以及两个串联重复的21碱基对序列元件,每个元件都定位于mRNA起始位点。每个起始区域重复序列都包含一个与ets相关转录因子的结合位点,该转录因子对特征性的GGAA ets序列基序具有特异性,并与ets结构域导向的单克隆泛ets抗体发生反应。这两个21碱基对的重复序列足以产生转录活性,表明与ets相关的因子可能参与转录激活和起始位点定位。通过比较用纯化的重组表达GABPα和β亚基形成的DNA - 蛋白质复合物的电泳迁移率和免疫反应性,发现COS核提取物中发现的与ets相关的蛋白质与GA结合蛋白(GABP)相同或密切相关。Sp1和与GABP相关的因子也与另一个小鼠细胞色素氧化酶亚基基因COXVb结合。这两个基因相似的启动子特征提示了此类呼吸蛋白之间可能存在的协同转录调控方式。