LINE-1-mediated retrotransposition in human pluripotent stem cells: Consequences for genomic stability of hES and hiPS cells and its derivatives

多能干细胞来源于发育中的囊胚的内部细胞群（人胚胎干细胞，hESCs）或由成纤维细胞重编程产生（人诱导多能干细胞，hiPSCs），在再生医学中具有巨大的治疗前景。然而，一个重要的悬而未决的问题是，这些细胞或它们的衍生物是否真的安全。就物种而言，目前尚不清楚多能干细胞在其产生、扩增和分化过程中是否保持了基因组的完整性。在扩增或分化过程中出现的基因突变可能会破坏干细胞治疗。这种突变可以由人类非ltr逆转录转座子（LINE1或L1、Alu、SVA）诱导，它们代表了目前被动员的人类内源性逆转录因子群。大约35%的人类基因组是由非ltr逆转录转座子动员的结果，这是由l1编码的蛋白质机器执行的。我们将研究hESCs和hiPSCs对L1反转录转位的支持是否不同，并分析L1反转录转位对这些细胞的基因组不稳定程度。我们想要确定L1活性在组织培养多能干细胞中经常观察到的核型异常形成中的作用，以及分化对L1动员率的影响。我们将评估在hESC和hiPSC细胞系中是否存在可能影响邻近基因表达的特定基因组区域的L1整合偏好。所提出的方法将有助于阐明l1介导的逆转录转位是否有助于hESCs和hiPSCs的基因组流动性和变异性，并将有助于评估多能干细胞中内源性移动元件的活性，以评估其治疗安全性。

The use of pluripotent stem cells derived from the inner cell mass of developing blastocysts (human embryonic stem cells, hESCs) or generated by reprogramming of fibroblasts (human induced pluripo-tent stem cells, hiPSCs) holds great therapeutic promise for regenerative medicine. However, a vital unanswered question is whether these cells or their derivatives are truly safe for administration. Spe-cifically, it is unclear whether the integrity of the genome of pluripotent stem cells is maintained during their generation, expansion and differentiation. The appearance of genetic mutations during their ex-pansion or differentiation could undermine stem cell therapies. Such mutations could be induced by human non-LTR retrotransposons (LINE1 or L1, Alu, SVA) which represent the currently mobilized group of human endogenous retroelements. About 35% of the human genome are the consequence of the mobilization of non-LTR retrotransposons which is executed by the L1-encoded protein machin-ery. We will investigate whether hESCs and hiPSCs differ in their support for L1 retrotransposition and analyze the extent of genomic destabilization of these cells by L1- retrotransposition. We want to determine both the role of L1 activity in the formation of karyotypic abnormalities that are frequently observed in pluripotent stem cells in tissue culture, and the effect of differentiation on L1 mobilization rates. We will evaluate if there are any L1 integration preferences for specific genomic regions in hESC and hiPSC lines that could potentially affect neighbouring gene expression. The proposed ap-proaches will help to elucidate whether L1-mediated retrotransposition can contribute to genome fluid-ity and variability of hESCs and hiPSCs, and will be useful to assess the activity of endogenous mobile elements in pluripotent stem cells in order to evaluate their safety for therapy.