Transcriptional regulation of rod photoreceptor homeostasis revealed by in vivo NRL targetome analysis.
Transcriptional regulation of rod photoreceptor homeostasis revealed by in vivo NRL targetome analysis.
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DOI：
10.1371/journal.pgen.1002649
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发表时间：
2012
期刊：
影响因子：
4.5
通讯作者：
Swaroop A
中科院分区：
文献类型：
--
作者：
Hao H;Kim DS;Klocke B;Johnson KR;Cui K;Gotoh N;Zang C;Gregorski J;Gieser L;Peng W;Fann Y;Seifert M;Zhao K;Swaroop A
严格控制稳态对于神经元的功能维持和存活至关重要。在哺乳动物视网膜中，基本基序亮氨酸拉链转录因子NRL决定视杆细胞与视锥细胞的命运，并激活许多视杆细胞特异性基因的表达。在这里，我们报告了NRL为中心的基因调控网络的综合分析，通过耦合染色质免疫沉淀，然后从Illumina和ABI平台的高通量测序（ChIP-Seq）数据与全球表达谱和体内敲除研究。我们鉴定了大约300个直接NRL靶基因。其中，22个NRL目标与人类视网膜营养不良相关，而95个映射到尚未克隆的视网膜疾病基因座的区域。NRL ChIP-Seq峰序列的计算机模拟分析揭示了不同转录因子结合位点集合的富集。具体地说，我们发现参与感光功能的基因包括NRL和同源结构域蛋白CRX的结合位点。对26个ChIP-Seq区域的评估验证了它们在报告基因测定中的增强子功能。在体内敲除16个NRL靶基因导致视杆细胞死亡或形态异常，表明它们在维持视网膜功能中的重要性。我们还确定了组蛋白去甲基化酶Kdm5b作为一个新的二级节点在NRL转录层次。外显子阵列分析的流式分选光感受器中，Kdm5b被敲低的shRNA表明其在调节杆表达的基因的作用。我们的研究确定视网膜营养不良的候选基因，定义光感受器表达基因的顺式调控模块，并提供了一个框架，用于解码转录调控网络，决定杆稳态。视网膜中的视杆和视锥光感受器是高度特化的神经元，其分别在昏暗和明亮的光下捕获光子。视杆细胞光感受器的丧失是大多数视网膜神经退行性疾病的早期临床表现，最终导致视锥细胞死亡和失明。转录因子NRL是视杆细胞命运和基因表达的关键调节因子。在这里，我们报告了NRL的全球转录靶点的综合分析。我们已经发现，NRL和CRX结合位点都存在于参与感光功能的基因中，这意味着它们之间存在密切的协同关系。对小鼠视网膜中16个NRL靶基因的体内功能丧失分析导致感光细胞死亡或形态异常。此外，我们确定组蛋白去甲基化酶Kdm5b作为NRL为中心的基因调控网络中的次要节点。我们的研究将NRL靶基因鉴定为视网膜退行性疾病患者突变筛查的优秀候选基因，它们为阐明视杆细胞稳态的调节和涉及光感受器功能障碍的疾病的治疗干预靶点提供了基础。
A stringent control of homeostasis is critical for functional maintenance and survival of neurons. In the mammalian retina, the basic motif leucine zipper transcription factor NRL determines rod versus cone photoreceptor cell fate and activates the expression of many rod-specific genes. Here, we report an integrated analysis of NRL-centered gene regulatory network by coupling chromatin immunoprecipitation followed by high-throughput sequencing (ChIP–Seq) data from Illumina and ABI platforms with global expression profiling and in vivo knockdown studies. We identified approximately 300 direct NRL target genes. Of these, 22 NRL targets are associated with human retinal dystrophies, whereas 95 mapped to regions of as yet uncloned retinal disease loci. In silico analysis of NRL ChIP–Seq peak sequences revealed an enrichment of distinct sets of transcription factor binding sites. Specifically, we discovered that genes involved in photoreceptor function include binding sites for both NRL and homeodomain protein CRX. Evaluation of 26 ChIP–Seq regions validated their enhancer functions in reporter assays. In vivo knockdown of 16 NRL target genes resulted in death or abnormal morphology of rod photoreceptors, suggesting their importance in maintaining retinal function. We also identified histone demethylase Kdm5b as a novel secondary node in NRL transcriptional hierarchy. Exon array analysis of flow-sorted photoreceptors in which Kdm5b was knocked down by shRNA indicated its role in regulating rod-expressed genes. Our studies identify candidate genes for retinal dystrophies, define cis-regulatory module(s) for photoreceptor-expressed genes and provide a framework for decoding transcriptional regulatory networks that dictate rod homeostasis. The rod and cone photoreceptors in the retina are highly specialized neurons that capture photons under dim and bright light, respectively. Loss of rod photoreceptors is an early clinical manifestation in most retinal neurodegenerative diseases that eventually result in cone cell death and blindness. The transcription factor NRL is a key regulator of rod photoreceptor cell fate and gene expression. Here, we report an integrated analysis of the global transcriptional targets of NRL. We have discovered that both NRL and CRX binding sites are present in genes involved in photoreceptor function, implying their close synergistic relationship. In vivo loss-of-function analysis of 16 NRL target genes in the mouse retina resulted in death or abnormal morphology of photoreceptor cells. Furthermore, we identified histone demethylase Kdm5b as a secondary node in the NRL-centered gene regulatory network. Our studies identify NRL target genes as excellent candidates for mutation screening of patients with retinal degenerative diseases, and they provide the foundation for elucidating regulation of rod homeostasis and targets for therapeutic intervention in diseases involving photoreceptor dysfunction.
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影响因子：
30.8
作者：
Hollander, MC;Sheikh, MS;Fornace, AJ
通讯作者：
Fornace, AJ
影响因子：
16.2
作者：
Chen, SM;Wang, QL;Zack, DJ
通讯作者：
Zack, DJ
DOI：
10.1073/pnas.0902425106
发表时间：
2009-10-13
影响因子：
11.1
作者：
Jia, Li;Oh, Edwin C. T.;Forrest, Douglas
通讯作者：
Forrest, Douglas
影响因子：
16
作者：
Fujiwara T;O'Geen H;Keles S;Blahnik K;Linnemann AK;Kang YA;Choi K;Farnham PJ;Bresnick EH
通讯作者：
Bresnick EH
影响因子：
3.5
作者：
Cheng, Hong;Aleman, Tomas S.;Swaroop, Anand
通讯作者：
Swaroop, Anand