CRISPRi-mediated functional analysis of NKX2-1-binding sites in the lung.
CRISPRi-mediated functional analysis of NKX2-1-binding sites in the lung.
复制标题
肺部 NKX2-1 结合位点的 CRISPRi 介导的功能分析。
DOI：
10.1038/s42003-021-02083-4
复制
发表时间：
2021-05-12
影响因子：
5.9
通讯作者：
Maeda Y
中科院分区：
文献类型：
--
作者：
Stuart WD;Fink-Baldauf IM;Tomoshige K;Guo M;Maeda Y
转录因子NKX 2 -1/TTF-1参与肺病理生理学，包括呼吸、先天防御和肿瘤发生。为了了解NKX 2 -1调节参与这种病理生理学的基因的机制，我们之前进行了ChIP-seq并鉴定了全基因组NKX 2 -1结合位点，这表明NKX 2 -1不仅与近端启动子区域结合，而且与NKX 2 -1调节的基因的多个基因内和基因间区域结合。然而，这些区域，特别是非近端区域，与NKX 2 -1结合的作用尚未确定。在这里，使用CRISPRi（CRISPR/dCas 9-KRAB），我们仔细检查了NKX 2 -1结合的19个区域/位点的功能作用，这些区域/位点位于参与呼吸和先天防御（SFTPB，LAMP 3，SFTPA 1，SFTPA 2）和肺肿瘤发生（MYBPH，LMO 3，CD 274/PD-L1）的基因中。值得注意的是，CRISPRi方法揭示了NKX 2 -1结合位点的一部分在功能上是不可或缺的，而其余部分对于基因的表达是不可或缺的，这表明NKX 2 -1结合位点的功能作用是不平等的。转录因子NKX 2 -1是肺病理生理学的关键调节因子，但其近端启动子区域外的结合位点的重要性尚不清楚。在这里，William Stuart和Iris Fink-Baldauf等人使用CRISPRi来询问19个NKX 2 -1结合位点，并确定对呼吸，先天免疫防御和肿瘤发生重要的特定位点。
The transcription factor NKX2-1/TTF-1 is involved in lung pathophysiology, including breathing, innate defense and tumorigenesis. To understand the mechanism by which NKX2-1 regulates genes involved in such pathophysiology, we have previously performed ChIP-seq and identified genome-wide NKX2-1-binding sites, which revealed that NKX2-1 binds to not only proximal promoter regions but also multiple intra- and inter-genic regions of the genes regulated by NKX2-1. However, the roles of such regions, especially non-proximal ones, bound by NKX2-1 have not yet been determined. Here, using CRISPRi (CRISPR/dCas9-KRAB), we scrutinize the functional roles of 19 regions/sites bound by NKX2-1, which are located in genes involved in breathing and innate defense (SFTPB, LAMP3, SFTPA1, SFTPA2) and lung tumorigenesis (MYBPH, LMO3, CD274/PD-L1). Notably, the CRISPRi approach reveals that a portion of NKX2-1-binding sites are functionally indispensable while the rest are dispensable for the expression of the genes, indicating that functional roles of NKX2-1-binding sites are unequally yoked. The transcription factor NKX2-1 is a key regulator of lung pathophysiology, but the importance of its binding sites outside of proximal promoter regions is unclear. Here, William Stuart and Iris Fink-Baldauf et al. use CRISPRi to interrogate 19 NKX2-1 binding sites and identify specific sites important for breathing, innate immune defense, and tumorigenesis.
登录
查看更多内容
影响因子：
10.5
作者：
Iwafuchi-Doi M;Zaret KS
通讯作者：
Zaret KS
影响因子：
--
作者：
Holzinger, A;Dingle, S;Whitsett, JA
通讯作者：
Whitsett, JA
影响因子：
9.3
作者：
Durand NC;Shamim MS;Machol I;Rao SS;Huntley MH;Lander ES;Aiden EL
通讯作者：
Aiden EL
影响因子：
44.1
作者：
通讯作者：
--
影响因子：
16
作者：
Heinz S;Benner C;Spann N;Bertolino E;Lin YC;Laslo P;Cheng JX;Murre C;Singh H;Glass CK
通讯作者：
Glass CK