Variegated gene expression caused by cell-specific long-range DNA interactions

Mammalian genomes contain numerous DNA elements with potential transcription regulatory function but unknown target genes. We used transgenic, gain-of-function mice with an ectopic copy of the beta-globin locus control region (LCR) to better understand how regulatory elements dynamically search the genome for target genes. We find that the LCR samples a restricted nuclear sub-volume in which it forms preferential contacts with genes controlled by shared transcription factors. One contacted gene, betah1, located on another chromosome, is upregulated, providing genetic demonstration that mammalian enhancers can function between chromosomes. Upregulation is not pan-cellular but confined to selected 'jackpot' cells significantly enriched for inter-chromosomal LCR-betah1 interactions. This implies that long-range DNA contacts are relatively stable and cell-specific and, when functional, cause variegated expression. We refer to this as spatial effect variegation (SEV). The data provide a dynamic and mechanistic framework for enhancer action, important for assigning function to the one- and three-dimensional structure of DNA. A knock-in mouse strain was compared to a wild-type FVB strain. The knock-in mouse strain carries a human beta-globin LCR in the Rad23a locus. We performed 4C for the Rad23a locus in the knock-in and the wild-type strain. Biological replicates were analyzed in a reversed dye orientation. At the RNA level these mice were characterized with Affymetrix expression arrays. We analyzed three biological replicates for the WT and the knock-in.哺乳动物基因组中存在大量具备潜在转录调控功能、但尚未明确其靶基因的DNA元件。本研究利用携带异位β-珠蛋白基因座控制区（LCR）拷贝的转基因功能获得型小鼠，旨在更深入解析调控元件如何动态搜索基因组以识别靶基因。研究发现，LCR会靶向限定的细胞核亚区室，在此区域内它优先与受共有转录因子调控的基因形成相互作用。其中一个发生相互作用的基因betah1位于另一条染色体上，其表达被上调，这从遗传学层面证实了哺乳动物增强子可跨染色体发挥功能。这种表达上调并非全细胞普遍存在，而是仅局限于特定的‘jackpot细胞’——这类细胞中跨染色体LCR-betah1相互作用的富集程度显著升高。这表明长距离DNA相互作用相对稳定且具有细胞特异性，当其发挥功能时会引发镶嵌式表达，我们将这一现象命名为空间效应镶嵌现象（SEV）。本研究数据为增强子作用机制提供了动态的分析框架，这对于解析DNA一维与三维结构的功能具有重要意义。 本研究将敲入小鼠品系与野生型FVB品系进行对比：敲入小鼠品系的Rad23a基因座上整合了人源β-珠蛋白LCR。我们分别对敲入品系与野生型品系的Rad23a基因座开展了4C实验，实验设置了反向染料标记的生物学重复进行分析。在RNA层面，我们通过Affymetrix表达芯片对这些小鼠进行了表达谱分析，其中野生型与敲入品系各设置了3个生物学重复样本。