Stanojevic Dusan, Young Richard A
Crosslink Genetics Corporation, One Kendall Square, Building 600, PMB 255, Cambridge, MA 02139, USA.
Biochemistry. 2002 Jun 11;41(23):7209-16. doi: 10.1021/bi015906b.
The use of synthetic chemical moieties to design fully functional analogues of transcription factors will give rise to novel molecular tools for targeted gene regulation. Here we demonstrate that a synthetic molecule based on a nonpeptidic DNA-binding domain can be engineered to function as a highly potent transcription factor in vitro and in an intracellular context. The structure of this artificial transcription factor (ATF) consists of three parts: (i) triple-helix-forming oligonucleotide as a DNA-binding domain; (ii) composite linker moiety; and (iii) short synthetic peptide. The direct comparison of ATFs with natural transcription factors in in vitro assays reveals the ability of ATFs to initiate RNA transcription at the correct initiation site. In addition, the transcriptional activation potency of ATFs in vitro matches or exceeds the potency of GAL4-VP16, one of the strongest natural transcriptional activators. This remarkable biological activity is explained as a function of ATF's chemical structure. We also demonstrate for the first time that ATFs possess substantial ability to activate transcription in tissue culture cells, thus opening a prospect for practical applications in basic and applied research. The specific molecular design employed in the synthesis of ATFs may lead to the development of novel gene-targeting pharmaceuticals for treatment of fatal and chronic diseases.
利用合成化学基团设计转录因子的全功能类似物将产生用于靶向基因调控的新型分子工具。在此，我们证明基于非肽DNA结合域的合成分子可被设计成在体外和细胞内环境中发挥高效转录因子的功能。这种人工转录因子（ATF）的结构由三部分组成：（i）作为DNA结合域的三链螺旋形成寡核苷酸；（ii）复合连接基团；（iii）短合成肽。在体外试验中对ATF与天然转录因子的直接比较揭示了ATF在正确起始位点启动RNA转录的能力。此外，ATF在体外的转录激活效力与最强的天然转录激活剂之一GAL4-VP16的效力相当或超过其效力。这种显著的生物学活性被解释为ATF化学结构的一种功能。我们还首次证明ATF在组织培养细胞中具有显著的激活转录的能力，从而为基础研究和应用研究中的实际应用开辟了前景。ATF合成中采用的特定分子设计可能会导致开发用于治疗致命和慢性疾病的新型基因靶向药物。
