Genome Toxicity and Impaired Stem Cell Function after Conditional Activation of CreER(T2) in the Intestine.
Genome Toxicity and Impaired Stem Cell Function after Conditional Activation of CreER(T2) in the Intestine.
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DOI：
10.1016/j.stemcr.2018.10.014
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发表时间：
2018-12-11
影响因子：
5.9
通讯作者：
Samuelson LC
中科院分区：
文献类型：
--
作者：
Bohin N;Carlson EA;Samuelson LC
使用他莫昔芬诱导的CreERT2系统，可以在完整动物中以细胞类型特异性的方式暂时控制遗传重组，从而允许解剖哺乳动物生理学的分子基础。在这里，我们提出了用于肠道干细胞分析的CreERT2技术的一个重大缺陷。使用肠道特异性Villin-CreERT2小鼠菌株，我们观察到辐照后肠道再生延迟。Villin-CreERT2激活与DNA损伤和隐性loxP位点切割有关。对干细胞特异性CreERT2菌株的分析表明，基因组毒性损害隐窝基柱状干细胞的功能，导致类器官启动活性丧失。重要的是，干细胞损伤是短暂的，在他莫昔芬治疗后7天恢复正常。我们的研究结果表明，利用CreERT2的小鼠遗传实验应该考虑由CreERT2激活导致的干细胞对基因组毒性的敏感性增强的混杂效应。CreERT2激活诱导小鼠肠干细胞（ISC）毒性ISC特异性CreERT2菌株激活后类器官形成受损绒毛蛋白-CreERT2小鼠的遗传毒性和隐窝再生受损ISC功能受损和遗传毒性在激活后7天修复Samuelson和他的同事证明小鼠肠道中CreERT2的激活会导致肠干细胞（ISC）毒性。ISC-CreERT2菌株的干细胞功能受损表现为类器官形成能力降低。此外，γ辐照诱导的ISC损伤后，绒毛蛋白- creert2小鼠的隐窝再生受损。隐性loxP位点切割导致的DNA损伤提示了ISC遗传毒性的机制。
With the tamoxifen-inducible CreERT2 system, genetic recombination can be temporally controlled in a cell-type-specific manner in intact animals, permitting dissection of the molecular underpinnings of mammalian physiology. Here we present a significant drawback to CreERT2 technology for analysis of intestinal stem cells. Using the intestine-specific Villin-CreERT2 mouse strain, we observed delayed intestinal regeneration post irradiation. Villin-CreERT2 activation was associated with DNA damage and cryptic loxP site cleavage. Analysis of stem cell-specific CreERT2 strains showed that the genome toxicity impairs function of crypt base columnar stem cells, resulting in loss of organoid initiating activity. Importantly, the stem cell impairment is short-lived, with return to normal by 7 days post tamoxifen treatment. Our findings demonstrate that mouse genetic experiments that utilize CreERT2 should consider the confounding effects of enhanced stem cell sensitivity to genome toxicity resulting from CreERT2 activation. Intestinal stem cell (ISC) toxicity induced in mice by CreERT2 activation Impaired organoid formation after activation of ISC-specific CreERT2 strains Genotoxicity and impaired crypt regeneration in Villin-CreERT2 mice Impaired ISC function and genotoxicity repaired by 7 days after activation Samuelson and colleagues demonstrate that activation of CreERT2 in the mouse intestine leads to intestinal stem cell (ISC) toxicity. Impaired stem cell function was shown by reduced organoid-forming ability in ISC-CreERT2 strains. Also, Villin-CreERT2 mice exhibited impaired crypt regeneration after ISC injury induced by γ-irradiation. DNA damage due to cryptic loxP site cleavage suggested a mechanism of ISC genotoxicity.
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