Synthesizing AND gate genetic circuits based on CRISPR-Cas9 for identification of bladder cancer cells
Synthesizing AND gate genetic circuits based on CRISPR-Cas9 for identification of bladder cancer cells
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DOI：
10.1038/ncomms6393
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发表时间：
2014-11-01
影响因子：
16.6
通讯作者：
Cai, Zhiming
中科院分区：
文献类型：
--
作者：
Liu, Yuchen;Zeng, Yayue;Cai, Zhiming
传统的癌症基因治疗策略对特异性和疗效的控制有限。克服这些限制的一种可能的方法是构建逻辑电路。本文提出了基于CRISPR-Cas9系统的模块化与门电路。电路整合来自两个启动子的细胞信息作为输入，只有当两个输入在被测试的细胞系中都激活时，才激活输出基因。使用荧光素酶报告基因作为输出基因，我们发现该回路特异性检测膀胱癌细胞，与人类端粒酶逆转录酶-肾细胞荧光素酶构建体相比，荧光素酶表达显著增强。我们还通过将输出替换为其他细胞功能基因（包括hBAX、p21和E-cadherin）来测试设计的模块化。该回路通过调控相应基因，有效抑制膀胱癌细胞生长，诱导细胞凋亡，降低细胞活力。该方法为体外靶向和控制膀胱癌细胞提供了一个合成生物学平台。
The conventional strategy for cancer gene therapy offers limited control of specificity and efficacy. A possible way to overcome these limitations is to construct logic circuits. Here we present modular AND gate circuits based on CRISPR-Cas9 system. The circuits integrate cellular information from two promoters as inputs and activate the output gene only when both inputs are active in the tested cell lines. Using the luciferase reporter as the output gene, we show that the circuit specifically detects bladder cancer cells and significantly enhances luciferase expression in comparison to the human telomerase reverse transcriptase-renilla luciferase construct. We also test the modularity of the design by replacing the output with other cellular functional genes including hBAX, p21 and E-cadherin. The circuits effectively inhibit bladder cancer cell growth, induce apoptosis and decrease cell motility by regulating the corresponding gene. This approach provides a synthetic biology platform for targeting and controlling bladder cancer cells in vitro.