Impeding transcription of expanded microsatellite repeats by deactivated Cas9
Impeding transcription of expanded microsatellite repeats by deactivated Cas9
复制标题
失活的 Cas9 阻碍微卫星重复序列的转录
DOI：
10.1101/185496
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发表时间：
2017
期刊：
影响因子：
--
通讯作者：
Eric T. Wang
中科院分区：
文献类型：
--
作者：
Belinda S. Pinto;Tanvi Saxena;Ruan Oliveira;H. Méndez;J. Cleary;Lance T. Denes;O. McConnell;Juan D. Arboleda;Guangbin Xia;M. Swanson;Eric T. Wang
扩增的微卫星重复序列的转录与多种人类疾病相关，包括强直性肌营养不良、Fuchs角膜内皮营养不良和C9 orf 72-ALS/FTD。消除或减少由这些扩展的基因座产生的RNA和蛋白质的产生具有治疗益处。在这里，我们测试了Cas9酶的失活形式阻碍跨扩展的微卫星转录的假设。我们观察到在将dCas 9直接靶向重复序列后，含有重复序列的RNA的丰度的重复长度依赖性、PAM依赖性和链依赖性降低。在DM 1细胞中挽救了异常剪接模式，并且DM 1、DM 2和C9 orf 72-ALS/FTD细胞特征性RAN肽的产生显著降低。通过腺相关病毒全身递送dCas 9/gRNA导致病理性RNA病灶的减少、氯离子通道1蛋白表达的拯救和肌强直的减少。这些观察结果表明，微卫星重复序列的RNA的转录比其他RNA的转录对扰动更敏感，这表明了潜在的可行的治疗干预策略。
Transcription of expanded microsatellite repeats is associated with multiple human diseases, including myotonic dystrophy, Fuchs’ endothelial corneal dystrophy, and C9orf72-ALS/FTD. Eliminating or reducing production of RNA and proteins arising from these expanded loci holds therapeutic benefit. Here, we tested the hypothesis that a deactivated form of the Cas9 enzyme impedes transcription across expanded microsatellites. We observed a repeat length-, PAM-, and strand-dependent reduction in the abundance of repeat-containing RNAs upon targeting dCas9 directly to repeat sequences. Aberrant splicing patterns were rescued in DM1 cells, and production of RAN peptides characteristic of DM1, DM2, and C9orf72-ALS/FTD cells was drastically decreased. Systemic delivery of dCas9/gRNA by adeno-associated virus led to reductions in pathological RNA foci, rescue of chloride channel 1 protein expression, and decreased myotonia. These observations suggest that transcription of microsatellite repeat-containing RNAs is more sensitive to perturbation than transcription of other RNAs, indicating potentially viable strategies for therapeutic intervention.
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DOI：
10.1073/pnas.0501507102
发表时间：
2005-04-19
影响因子：
11.1
作者：
Harper, SQ;Staber, PD;Davidson, BL
通讯作者：
Davidson, BL
DM1 诱导多能干细胞产生神经细胞，作为研究中枢神经系统神经发病机制的细胞模型。
影响因子：
1.6
作者：
Xia,Guangbin;Santostefano,KatherineE;Goodwin,Marianne;Liu,Jilin;Subramony,SH;Swanson,MauriceS;Terada,Naohiro;Ashizawa,Tetsuo
通讯作者：
Ashizawa,Tetsuo
影响因子：
1.2
作者：
Lampe, Sara E. Gombash;Kaspar, Brian K.;Foust, Kevin D.
通讯作者：
Foust, Kevin D.
影响因子：
56.9
作者：
Philips, AV;Timchenko, LT;Cooper, TA
通讯作者：
Cooper, TA
影响因子：
4.8
作者：
Zolotukhin, S;Potter, M;Snyder, RO
通讯作者：
Snyder, RO