Valerio D, van der Putten H, Botteri F M, Hoogerbrugge P M
Radiobiological Institute TNO, Rijswijk, The Netherlands.
Nucleic Acids Res. 1988 Nov 11;16(21):10083-97. doi: 10.1093/nar/16.21.10083.
The promoter of the human gene for adenosine deaminase (ADA) is extremely G/C-rich, contains several G/C-box motifs (GGGCGGG) and lacks any apparent TATA or CAAT boxes. These features are commonly found in promoters of genes that lack a strong tissue specificity, and are referred to as "housekeeping genes". Like other housekeeping genes, the ADA gene is expressed in all tissues. However, there is a considerable variation in the levels of expression of the ADA protein in different tissues. In order to study the activity of the ADA promoter, transgenic mice were generated that harbor a chimeric gene composed of the ADA promoter linked to a reporter gene encoding the bacterial enzyme Chloramphenicol Acetyl Transferase (CAT). These mice reproducibly showed CAT expression in all tissues examined, including the hemopoietic organs (spleen, thymus and bone marrow). However, examination of the actual cell types expressing the CAT gene revealed the ADA promoter to be inactive in the hemopoietic cells. This was substantiated by a transplantation experiment in which bone marrow from ADA-CAT transgenic mice was used to reconstitute the hemopoietic compartment of lethally irradiated mice. The engrafted recipients revealed strongly reduced CAT activity in their hemopoietic organs. The lack of expression in hemopoietic cells was further shown to be correlated with a hypermethylated state of the transgene. Combined, our data suggest that the ADA promoter sequences tested can direct expression in a wide variety of tissues as expected for a regular housekeeping gene promoter. However, the activity of the ADA promoter fragment did not reflect the tissue-specific variations in expression levels of the endogenous ADA gene. Additionally, regulatory elements are needed for expression in the hemopoietic cells.
人类腺苷脱氨酶（ADA）基因的启动子富含G/C，包含几个G/C盒基序（GGGCGGG），且没有明显的TATA盒或CAAT盒。这些特征常见于缺乏强组织特异性的基因启动子中，这类基因被称为“管家基因”。与其他管家基因一样，ADA基因在所有组织中均有表达。然而，不同组织中ADA蛋白的表达水平存在相当大的差异。为了研究ADA启动子的活性，构建了转基因小鼠，其携带一个嵌合基因，该基因由与编码细菌酶氯霉素乙酰转移酶（CAT）的报告基因相连的ADA启动子组成。这些小鼠在所有检测的组织中均可重复性地显示出CAT表达，包括造血器官（脾脏、胸腺和骨髓）。然而，对实际表达CAT基因的细胞类型进行检测发现，ADA启动子在造血细胞中无活性。这一点在一项移植实验中得到了证实，该实验使用ADA-CAT转基因小鼠的骨髓来重建受致死性照射小鼠的造血系统。移植后的受体在其造血器官中显示出CAT活性大幅降低。造血细胞中缺乏表达进一步表明与转基因的高甲基化状态相关。综合来看，我们的数据表明，所测试的ADA启动子序列能够如常规管家基因启动子预期的那样，在多种组织中指导基因表达。然而，ADA启动子片段的活性并未反映内源性ADA基因表达水平的组织特异性差异。此外，造血细胞中的表达还需要调控元件。
