A new approach to transcription factor screening for reprogramming of fibroblasts to cardiomyocyte-like cells
A new approach to transcription factor screening for reprogramming of fibroblasts to cardiomyocyte-like cells
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DOI：
10.1016/j.yjmcc.2012.04.010
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发表时间：
2012-09-01
影响因子：
5
通讯作者：
Ravens, Ursula
中科院分区：
文献类型：
--
作者：
Protze, Stephanie;Khattak, Shahryar;Ravens, Ursula
几种转录因子的同时过表达已经成为一种成功的策略，可以将成纤维细胞转化为其他类型的细胞，包括多能细胞、神经元和心肌细胞。因子的选择和筛选是至关重要的，通常涉及检测大量转录因子，然后连续移除单个因子。在这里，为了确定促进小鼠成纤维细胞重编程为心肌细胞的心脏转录因子组合，我们直接筛选了所有由10个候选因子组成的三联体组合，并通过Q-PCR方法报告了多个心脏特异基因的诱导。通过这种筛选方法，Tbx5、MEF2C和MYOCD的组合被鉴定为与Tbx5的组合相比，可以上调更广泛的心脏基因谱。MEF2C和最近被证明诱导成纤维细胞重编程为心肌细胞的GATA4。Tbx5、MEF2C、MYOCD共转导的细胞表达心肌收缩蛋白，并可诱发出心肌样钾钠电流和动作电位。综上所述，本文提出的替代筛选方法避免了在复杂的转录因子混合物中掩盖其效力的转录因子的消除。此外，我们的结果指出了在评估重新编程时验证多个谱系特定基因的重要性。(C)2012爱思唯尔有限公司。保留所有权利。
The simultaneous overexpression of several transcription factors has emerged as a successful strategy to convert fibroblasts into other cell types including pluripotent cells, neurons, and cardiomyocytes. The selection and screening of factors are critical, and have often involved testing a large pool of transcription factors, followed by successive removal of single factors. Here, to identify a cardiac transcription factor combination facilitating mouse fibroblast reprogramming into cardiomyocytes, we directly screened all triplet combinations of 10 candidate factors combined with a Q-PCR assay reporting induction of multiple cardiac-specific genes. Through this screening method the combination of Tbx5, Mef2c, and Myocd was identified to upregulate a broader spectrum of cardiac genes compared to the combination of Tbx5. Mef2c, and Gata4 that was recently shown to induce reprogramming of fibroblasts into cardiomyocytes. Cells cotransduced with Tbx5, Mef2c, Myocd expressed cardiac contractile proteins, had cardiac-like potassium and sodium currents and action potentials could be elicited. In summary the alternative screening approach that is presented here avoided the elimination of transcription factors whose potency is masked in complex transcription factor mixes. Furthermore, our results point to the importance of verifying multiple lineage specific genes when assessing reprogramming. (C) 2012 Elsevier Ltd. All rights reserved.