Defective Cohesin in CdLS Mediates Gene Expression with Characteristics of Transcription Factor and Insulator Activity. Homo sapiens

The Cohesin apparatus has a canonical role in sister chromatid cohesion. Heterozygous mutations in Nipped B-like (NIPBL), SMC1A, and SMC3 have been found in 60% of probands with Cornelia de Lange Syndrome (CdLS), a dominant multi-system genetic disorder with variable expression. We have performed a genome-wide transcription assessment as well as cohesin binding analysis using human lymphoblastoid cell lines (LCLs) from probands with CdLS and controls. Here, we report a unique profile of genes dysregulated in CdLS that correlates with different clinical presentations. Genome-wide analysis of cohesin binding demonstrates a preference for intergenic regions suggesting a cis-regulatory function mimicking that of an insulator. However, the binding sites are enriched within the promoter regions of the dysregulated genes and are significantly decreased in CdLS probands, indicating an alternative role of cohesin as a classic transcription factor. Cohesin also co-localizes with CTCF at the boundary elements affecting neighboring gene expression in CdLS probands. We propose that the CdLS phenotype is the result of dysregulated gene expression rather than defective sister chromatid cohesion. Phenotype specific expression profiles are also described. Keywords: Disease state analysis, Genetic modification Overall design: To identify differentially expressed genes between CdLS patients and controls, age and gender matched samples from 16 normal Caucasian controls and 17 clinical severely affected Caucasian patients with NIPBL protein truncating mutations (nonsense or frameshift) were chosen as the training set for the discriminate analysis. To validate the expression pattern obtained from the training set, 6 samples including 1 healthy control, 1 Egyptian CdLS patient, 2 Roberts syndrome patients, and 2 Alagille patients were used as the testing set. All the 39 cell lines were growing anonymously and the processing of these 39 cell lines were randomized by genotypes to eliminate batch effects that may contribute to genotype-specific gene expression

黏连蛋白（Cohesin）复合物在姐妹染色单体黏连（sister chromatid cohesion）中发挥经典功能。研究发现，60%的科妮莉亚·德·朗格综合征（Cornelia de Lange Syndrome, CdLS）先证者携带Nipped B样蛋白（NIPBL）、SMC1A及SMC3的杂合突变；该疾病为一种表型可变的显性多系统遗传性疾病。本研究针对携带CdLS的先证者与健康对照的人类淋巴母细胞系（lymphoblastoid cell lines, LCLs），开展了全基因组转录分析及黏连蛋白结合位点检测。本研究报道了CdLS患者中失调基因的独特表达谱，其与不同临床表型存在相关性。全基因组黏连蛋白结合分析显示，黏连蛋白偏好结合基因间区域，提示其可发挥类似绝缘子（insulator）的顺式调控功能。但进一步分析发现，失调基因的启动子区域显著富集黏连蛋白结合位点，且此类结合在CdLS先证者中显著减少，表明黏连蛋白可作为经典转录因子发挥另一重功能。在CdLS先证者中，黏连蛋白还与CTCF共定位在影响邻近基因表达的边界元件处。我们提出，CdLS的表型源于基因表达失调而非姐妹染色单体黏连功能缺陷。本研究同时描述了表型特异性的表达谱。
关键词：疾病状态分析、遗传修饰
整体实验设计：为鉴定CdLS患者与健康对照间的差异表达基因，我们选取16例年龄、性别匹配的高加索健康对照，以及17例携带NIPBL蛋白截短突变（无义突变或移码突变）的高加索重度受累CdLS先证者样本，作为判别分析的训练集。为验证训练集得到的表达模式，我们选取6份样本作为测试集，包含1例健康对照、1例埃及CdLS患者、2例罗伯茨综合征（Roberts syndrome）患者及2例阿拉吉尔综合征（Alagille syndrome）患者。全部39株细胞系均以匿名方式培养，且我们按基因型对39株细胞系的处理流程进行随机化，以消除可能导致基因型特异性基因表达差异的批次效应（batch effects）。