Simultaneous single-cell three-dimensional genome and gene expression profiling Uncovers Dynamic Enhancer Connectivity Underlying Olfactory Receptor Choice [olfactory, single-cell RNA & Hi-C]

The simultaneous measurement of three-dimensional (3D) genome structure and gene expression of individual cells is critical for understanding our genome's structure-function relation, yet is extremely challenging for existing methods. Here we present Linking mRNA to Chromatin Architecture (LiMCA), which jointly profiles 3D genome and transcriptome with exceptional sensitivity and from low-input materials. Combining LiMCA and our high-resolution scATAC-seq assay, METATAC, we profiled the developing mouse olfactory epithelium and reported the first chromatin accessibility profiles and the first paired 3D genome structures and gene expression information of single neurons. We greatly expanded the repertoire of known OR enhancers, and discovered unexpected rules of their dynamics: ORs and their enhancers are most accessible during early differentiation, and the active OR typically does not associate with the largest enhancer hub. These findings offer valuable insights into how 3D connectivity of ORs and enhancers dynamically orchestrate the "one neuron-one receptor" selection process. simultaneous 3D genome and gene expression within the same single cell by LiMCA Each cell was named according to the developmental stage/age and mouse strain information. For example, "p007" indicates that the developmental stage/age of the mouse from which this neuron was derived is postnatal day 7. "bc" and "bdf1" indicate the mouse strain. We labeled F1 hybrids of C57BL/6J (female) and DBA/2J (male) as bdf1 and F1 hybrids of C57BL/6J (female) and CAST/EiJ (male) as bc.同步测定单个细胞的三维（3D）基因组结构与基因表达，对于解析基因组的结构-功能关联至关重要，但现有方法仍面临极大挑战。本研究介绍了mRNA-染色质构象连接技术（Linking mRNA to Chromatin Architecture, LiMCA），该方法可从低起始量样本中以优异灵敏度同时表征3D基因组与转录组。结合LiMCA与本团队开发的高分辨率单细胞转座酶可及性测序（single-cell Assay for Transposase-Accessible Chromatin using sequencing, scATAC-seq）实验体系METATAC，我们对发育中的小鼠嗅觉上皮进行了组学表征，首次获取了单个神经元的染色质可及性图谱、配对的3D基因组结构与基因表达信息。本研究大幅扩充了已知嗅觉受体（olfactory receptor, OR）增强子的集合，并揭示了其动态调控的意外规律：嗅觉受体与其增强子在早期分化阶段的可及性最高，且活跃的嗅觉受体通常并不与最大的增强子枢纽相关联。上述发现为理解嗅觉受体与增强子的三维连接如何动态调控“一个神经元-一个受体”的选择过程提供了极具价值的见解。通过LiMCA获取的同一单个细胞的同步3D基因组与基因表达数据集：每个细胞的命名依据其发育阶段/年龄与小鼠品系信息。例如，“p007”表示该神经元取自出生后第7天的小鼠；“bc”与“bdf1”代表小鼠品系：我们将C57BL/6J（雌性）与DBA/2J（雄性）的F1杂交子代命名为bdf1，将C57BL/6J（雌性）与CAST/EiJ（雄性）的F1杂交子代命名为bc。