Simple and effective generation of transgene-free induced pluripotent stem cells using an auto-erasable Sendai virus vector responding to microRNA-302
Simple and effective generation of transgene-free induced pluripotent stem cells using an auto-erasable Sendai virus vector responding to microRNA-302
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DOI：
10.1016/j.scr.2017.06.011
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发表时间：
2017-08-01
影响因子：
1.2
通讯作者：
Nakanishi, Mahito
中科院分区：
文献类型：
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作者：
Nishimura, Ken;Ohtaka, Manami;Nakanishi, Mahito
无转基因诱导多能干细胞（iPSC）在基础研究和潜在的临床应用方面都很有价值。我们先前报道了一种携带四种重编程因子（SeVdp-iPS）的复制缺陷型和持续性仙台病毒（SeVdp）载体可以有效地诱导产生无转基因的iPSCs。该载体可以稳定地同时表达所有四种因子而不需要染色体整合，并且可以通过抑制载体衍生的RNA依赖性RNA聚合酶而从重编程细胞中完全消除。在这里，我们描述了一种改进的SeVdp-iPS载体（SeVdp（KOSM）302 L），其响应于在多能干细胞（PSC）中独特表达的microRNA-302（miR-302）而自动擦除。SeVdp-302 L载体的基因表达和基因组复制在PSC中被强烈抑制，所述SeVdp-302 L载体在L mRNA的3'非翻译区含有miRNA-302 a靶序列。因此，SeVdp（KOSM）302 L诱导体细胞中重编程因子的表达，同时其从成功重编程以表达miR-302的细胞中自动擦除。由于这种载体可以在没有外源性短干扰RNA（siRNA）的帮助下将体细胞重编程为无转基因的iPSCs，因此我们在这里提出的结果表明，这种载体可能成为用于未来临床应用的人类iPSCs生成的宝贵工具。(C)2017作者出版社：Elsevier B.V
Transgene-free induced pluripotent stem cells (iPSCs) are valuable for both basic research and potential clinical applications. We previously reported that a replication-defective and persistent Sendai virus (SeVdp) vector harboring four reprogramming factors (SeVdp-iPS) can efficiently induce generation of transgene-free iPSCs. This vector can express all four factors stably and simultaneously without chromosomal integration and can be eliminated completely from reprogrammed cells by suppressing vector-derived RNA-dependent RNA polymerase. Here, we describe an improved SeVdp-iPS vector (SeVdp(KOSM) 302L) that is automatically erased in response to microRNA-302 (miR-302), uniquely expressed in pluripotent stem cells (PSCs). Gene expression and genome replication of the SeVdp-302L vector, which contains miRNA-302a target sequences at the 3' untranslated region of L mRNA, are strongly suppressed in PSCs. Consequently, SeVdp(KOSM) 302L induces expression of reprogramming factors in somatic cells, while it is automatically erased from cells successfully reprogrammed to express miR-302. As this vector can reprogram somatic cells into transgene-free iPSCs without the aid of exogenous short interfering RNA (siRNA), the results we present here demonstrate that this vector may become an invaluable tool for the generation of human iPSCs for future clinical applications. (C) 2017 The Authors. Published by Elsevier B.V