Expression of linear and novel circular forms of an INK4/ARF-associated non-coding RNA correlates with atherosclerosis risk.
Expression of linear and novel circular forms of an INK4/ARF-associated non-coding RNA correlates with atherosclerosis risk.
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DOI：
10.1371/journal.pgen.1001233
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发表时间：
2010-12-02
期刊：
影响因子：
4.5
通讯作者：
Sharpless NE
中科院分区：
文献类型：
--
作者：
Burd CE;Jeck WR;Liu Y;Sanoff HK;Wang Z;Sharpless NE
人类全基因组关联研究已将染色体9p21.3上INK 4/ARF（CDKN 2a/B）位点附近的单核苷酸多态性（SNP）与动脉粥样硬化性血管疾病（ASVD）的易感性联系起来。尽管该基因座编码三种特征良好的肿瘤抑制因子p16 INK 4a、p15 INK 4 b和ARF，但与ASVD最密切相关的SNP距离称为ANRIL（CDKN 2BAS）的长非编码RNA（ncRNA）内最近的编码基因约120 kb。虽然动脉粥样硬化风险等位基因纯合子个体显示ANRIL和编码INK 4/ARF转录物的表达降低，但这种远缘遗传变异影响INK 4/ARF表达的机制尚不清楚。在这里，使用cDNA末端快速扩增（RACE）和下一代RNA测序数据集的分析，我们确定了多个ANRIL物种的结构和丰度。这些物种中的每一个都以非常低的拷贝数存在于原代和培养细胞中;然而，只有含有INK 4/ARF基因座近端外显子的ANRIL亚型的表达与ASVD风险等位基因相关。令人惊讶的是，RACE还鉴定了含有非共线ANRIL外显子序列的转录物，其表达也与基因型和INK 4/ARF表达相关。这些非多聚腺苷酸化的RNA抵抗RNA酶R消化，并且可以使用面向外的引物进行PCR扩增，这表明它们代表可能由mRNA剪接的副产物产生的环状RNA结构。下一代DNA测序和剪接预测算法确定了ASVD风险区间内可能调节ANRIL剪接和环状ANRIL（cANRIL）产生的多态性。这些结果鉴定了源自ANRIL基因座的新型环状RNA产物，并表明9p21.3的致病变体通过调节ANRIL表达和/或结构来调节INK 4/ARF表达和ASVD风险。对人类基因组的无偏倚研究已经确定了染色体9p21.3上动脉粥样硬化性血管疾病（ASVD）的强遗传决定因素。该基因组区域不编码先前与ASVD相关的基因，但确实包含INK 4/ARF肿瘤抑制基因座。INK 4/ARF基因座的产物调节细胞分裂，这一过程被认为在ASVD病理学中很重要。我们和其他人认为9p21.3的遗传变异影响INK 4/ARF基因表达;然而，这些远距离多态性（> 100，000 bp）影响该位点转录的机制尚不清楚。ASVD相关的遗传变异位于称为ANRIL的非编码RNA（ncRNA）的预测结构内。基于最近的工作表明其他ncRNA可以抑制附近的编码基因，我们认为ANRIL结构可能调控INK 4/ARF基因表达。将分子分析与来自正常人类供体和癌细胞的各种细胞类型中的最先进的测序技术相结合，我们发现ANRIL编码稀有RNA转录物的异质物种。此外，我们鉴定了新的环状ANRIL亚型（cANRIL），其表达与INK 4/ARF转录和ASVD风险相关。这些研究提示了ANRIL结构影响INK 4/ARF表达和动脉粥样硬化易感性的新模型。
Human genome-wide association studies have linked single nucleotide polymorphisms (SNPs) on chromosome 9p21.3 near the INK4/ARF (CDKN2a/b) locus with susceptibility to atherosclerotic vascular disease (ASVD). Although this locus encodes three well-characterized tumor suppressors, p16INK4a, p15INK4b, and ARF, the SNPs most strongly associated with ASVD are ∼120 kb from the nearest coding gene within a long non-coding RNA (ncRNA) known as ANRIL (CDKN2BAS). While individuals homozygous for the atherosclerotic risk allele show decreased expression of ANRIL and the coding INK4/ARF transcripts, the mechanism by which such distant genetic variants influence INK4/ARF expression is unknown. Here, using rapid amplification of cDNA ends (RACE) and analysis of next-generation RNA sequencing datasets, we determined the structure and abundance of multiple ANRIL species. Each of these species was present at very low copy numbers in primary and cultured cells; however, only the expression of ANRIL isoforms containing exons proximal to the INK4/ARF locus correlated with the ASVD risk alleles. Surprisingly, RACE also identified transcripts containing non-colinear ANRIL exonic sequences, whose expression also correlated with genotype and INK4/ARF expression. These non-polyadenylated RNAs resisted RNAse R digestion and could be PCR amplified using outward-facing primers, suggesting they represent circular RNA structures that could arise from by-products of mRNA splicing. Next-generation DNA sequencing and splice prediction algorithms identified polymorphisms within the ASVD risk interval that may regulate ANRIL splicing and circular ANRIL (cANRIL) production. These results identify novel circular RNA products emanating from the ANRIL locus and suggest causal variants at 9p21.3 regulate INK4/ARF expression and ASVD risk by modulating ANRIL expression and/or structure. Unbiased studies of the human genome have identified strong genetic determinants of atherosclerotic vascular disease (ASVD) on chromosome 9p21.3. This region of the genome does not encode genes previously linked to ASVD, but does contain the INK4/ARF tumor suppressor locus. Products of the INK4/ARF locus regulate cell division, a process thought to be important in ASVD pathology. We and others have suggested that genetic variants in 9p21.3 influence INK4/ARF gene expression; however, the mechanisms by which these distant polymorphisms (>100,000 bp away) influence transcription of the locus is unknown. The ASVD–associated genetic variants lie within the predicted structure of a non-coding RNA (ncRNA) called ANRIL. Based upon recent work suggesting that other ncRNAs can repress nearby coding genes, we considered the possibility that ANRIL structure may regulate INK4/ARF gene expression. Coupling molecular analysis with state-of-the-art sequencing technologies in a wide variety of cell types from normal human donors and cancer cells, we found that ANRIL encodes a heterogeneous species of rare RNA transcripts. Moreover, we identified novel, circular ANRIL isoforms (cANRIL) whose expression correlated with INK4/ARF transcription and ASVD risk. These studies suggest a new model wherein ANRIL structure influences INK4/ARF expression and susceptibility to atherosclerosis.
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影响因子：
64.8
作者：
Frazer, Kelly A.;Ballinger, Dennis G.;Cox, David R.;Hinds, David A.;Stuve, Laura L.;Gibbs, Richard A.;Belmont, John W.;Boudreau, Andrew;Hardenbol, Paul;Leal, Suzanne M.;Pasternak, Shiran;Wheeler, David A.;Willis, Thomas D.;Yu, Fuli;Yang, Huanming;Zeng, Changqing;Gao, Yang;Hu, Haoran;Hu, Weitao;Li, Chaohua;Lin, Wei;Liu, Siqi;Pan, Hao;Tang, Xiaoli;Wang, Jian;Wang, Wei;Yu, Jun;Zhang, Bo;Zhang, Qingrun;Zhao, Hongbin;Zhao, Hui;Zhou, Jun;Gabriel, Stacey B.;Barry, Rachel;Blumenstiel, Brendan;Camargo, Amy;Defelice, Matthew;Faggart, Maura;Goyette, Mary;Gupta, Supriya;Moore, Jamie;Nguyen, Huy;Onofrio, Robert C.;Parkin, Melissa;Roy, Jessica;Stahl, Erich;Winchester, Ellen;Ziaugra, Liuda;Altshuler, David;Shen, Yan;Yao, Zhijian;Huang, Wei;Chu, Xun;He, Yungang;Jin, Li;Liu, Yangfan;Shen, Yayun;Sun, Weiwei;Wang, Haifeng;Wang, Yi;Wang, Ying;Xiong, Xiaoyan;Xu, Liang;Waye, Mary M. Y.;Tsui, Stephen K. W.;Wong, J. Tze-Fei;Galver, Luana M.;Fan, Jian-Bing;Gunderson, Kevin;Murray, Sarah S.;Oliphant, Arnold R.;Chee, Mark S.;Montpetit, Alexandre;Chagnon, Fanny;Ferretti, Vincent;Leboeuf, Martin;Olivier, Jean-Franccois;Phillips, Michael S.;Roumy, Stephanie;Sallee, Clementine;Verner, Andrei;Hudson, Thomas J.;Kwok, Pui-Yan;Cai, Dongmei;Koboldt, Daniel C.;Miller, Raymond D.;Pawlikowska, Ludmila;Taillon-Miller, Patricia;Xiao, Ming;Tsui, Lap-Chee;Mak, William;Song, You Qiang;Tam, Paul K. H.;Nakamura, Yusuke;Kawaguchi, Takahisa;Kitamoto, Takuya;Morizono, Takashi;Nagashima, Atsushi;Ohnishi, Yozo;Sekine, Akihiro;Tanaka, Toshihiro;Tsunoda, Tatsuhiko;Deloukas, Panos;Bird, Christine P.;Delgado, Marcos;Dermitzakis, Emmanouil T.;Gwilliam, Rhian;Hunt, Sarah;Morrison, Jonathan;Powell, Don;Stranger, Barbara E.;Whittaker, Pamela;Bentley, David R.;Daly, Mark J.;de Bakker, Paul I. W.;Barrett, Jeff;Chretien, Yves R.;Maller, Julian;McCarroll, Steve;Patterson, Nick;Pe'er, Itsik;Price, Alkes;Purcell, Shaun;Richter, Daniel J.;Sabeti, Pardis;Saxena, Richa;Schaffner, Stephen F.;Sham, Pak C.;Varilly, Patrick;Altshuler, David;Stein, Lincoln D.;Krishnan, Lalitha;Smith, Albert Vernon;Tello-Ruiz, Marcela K.;Thorisson, Gudmundur A.;Chakravarti, Aravinda;Chen, Peter E.;Cutler, David J.;Kashuk, Carl S.;Lin, Shin;Abecasis, Goncalo R.;Guan, Weihua;Li, Yun;Munro, Heather M.;Qin, Zhaohui Steve;Thomas, Daryl J.;McVean, Gilean;Auton, Adam;Bottolo, Leonardo;Cardin, Niall;Eyheramendy, Susana;Freeman, Colin;Marchini, Jonathan;Myers, Simon;Spencer, Chris;Stephens, Matthew;Donnelly, Peter;Cardon, Lon R.;Clarke, Geraldine;Evans, David M.;Morris, Andrew P.;Weir, Bruce S.;Tsunoda, Tatsuhiko;Johnson, Todd A.;Mullikin, James C.;Sherry, Stephen T.;Feolo, Michael;Skol, Andrew
通讯作者：
Skol, Andrew
影响因子：
64.8
作者：
通讯作者：
--
影响因子：
30.8
作者：
Bei, Jin-Xin;Li, Yi;Zeng, Yi-Xin
通讯作者：
Zeng, Yi-Xin
影响因子：
24
作者：
Gonzalez-Navarro, Herminia;Abu Nabah, Yafa Naim;Andres, Vicente
通讯作者：
Andres, Vicente
影响因子：
2.6
作者：
Emanuele, Enzo;Fontana, Jacopo M.;Geroldi, Diego
通讯作者：
Geroldi, Diego