Accurately quantifying the functional consequences of noncoding mosaic variants requires the pairing of DNA sequences with both accessible and closed chromatin architectures along individual DNA molecules—a pairing that cannot be achieved using traditional fragmentation-based chromatin assays. We demonstrate that targeted single-molecule chromatin fiber sequencing (Fiber-seq) achieves this, permitting single-molecule, long-read genomic, and epigenomic profiling across targeted >100 kb loci with ∼10-fold enrichment over untargeted sequencing. Targeted Fiber-seq reveals that pathogenic expansions of the DMPK CTG repeat that underlie Myotonic Dystrophy 1 are characterized by somatic instability and disruption of multiple nearby regulatory elements, both of which are repeat length-dependent. Furthermore, we reveal that therapeutic adenine base editing of the segmentally duplicated γ-globin (HBG1/HBG2) promoters in primary human hematopoietic cells induced toward an erythroblast lineage increases the accessibility of the HBG1 promoter as well as neighboring regulatory elements. Overall, we find that these non–protein coding mosaic variants can have complex impacts on chromatin architectures, including extending beyond the regulatory element harboring the variant.

中文翻译：

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使用靶向 Fiber-seq 解决镶嵌变异对染色质的影响


准确定量非编码嵌合变体的功能后果需要将 DNA 序列与单个 DNA 分子上可接近和封闭的染色质结构配对，而这种配对是使用传统的基于片段化的染色质分析无法实现的。我们证明靶向单分子染色质纤维测序 （Fiber-seq） 可以实现这一目标，允许对靶向 >100 kb 基因座进行单分子、长读长基因组和表观基因组分析，比非靶向测序富集 ∼ 10 倍。靶向 Fiber-seq 显示，强直性肌营养不良症 1 基础的 DMPK CTG 重复序列的致病性扩增的特征是体细胞不稳定和附近多个调节元件的破坏，这两者都是重复长度依赖性的。此外，我们揭示了原代人造血细胞中节段复制的 γ-珠蛋白 （HBG1/HBG2） 启动子的治疗性腺嘌呤碱基编辑诱导至成红细胞谱系，增加了 HBG1 启动子以及邻近调节元件的可及性。总体而言，我们发现这些非蛋白质编码马赛克变体会对染色质结构产生复杂的影响，包括延伸到包含该变体的调节元件之外。