G-quadruplex profiling in complex tissues using single-cell CUT&Tag

G-quadruplexes (G4) are non-canonical DNA structures that gained increasing attention for their potential roles in gene regulation, with implications in neurodegenerative diseases and cancer. Despite their biological significance, G4 structures have not been studied systematically across tissues and cell types. In this study, we employ G4 single-cell CUT&Tag (G4 scCUT&Tag) to characterize G4 landscapes in postnatal mouse brain cells, leveraging single-cell analytical approaches commonly used in scRNA-Seq and scATAC-Seq datasets. Using conventional single-cell omics workflows to process and explore our data, we distinguished different cell types based on G4 heterogeneity. Furthermore, we performed uncoupled multi-omics integration of G4 scCUT&Tag data with scRNA-Seq gene expression profiles, using both a covariance-based technique (canonical correlation analysis) and a transfer learning-based deep learning approach. These integrations not only revealed significant co-enrichment of G4 and gene expression signals, but demonstrated that G4 scCUT&Tag enables detailed examination of G4 heterogeneity in complex tissues and supports integrative analysis of G4 profiles with other omics layers, offering new insights into the epigenomic landscapes of the developing central nervous system. mouse ESC and NIH-3T3 were subjected to bulk and single cell G4 CUT&Tag (two biological replicates), and mouse brain were subjected to single cell G4 CUT&Tag

G-四链体（G-quadruplexes, G4）是一类非经典DNA结构，因其在基因调控中的潜在作用而受到日益广泛的关注，且与神经退行性疾病和癌症存在密切关联。尽管G4结构具有重要的生物学意义，但目前尚未在跨组织与细胞类型层面开展系统性研究。本研究采用G4单细胞CUT&Tag（G4 scCUT&Tag）技术，结合单细胞RNA测序（scRNA-Seq）与单细胞转座酶可及性测序（scATAC-Seq）中常用的单细胞分析方法，对出生后小鼠脑细胞的G4图谱进行表征。研究团队借助常规单细胞组学分析流程处理并解析所得数据，基于G4异质性区分出不同的细胞类型。此外，本研究还通过基于协方差的技术（典型相关分析）与基于迁移学习的深度学习方法，实现了G4 scCUT&Tag数据与scRNA-Seq基因表达谱的非耦合多组学整合。这些整合分析不仅揭示了G4与基因表达信号的显著共富集现象，同时证实G4 scCUT&Tag技术可在复杂组织中详细解析G4异质性，并支持G4图谱与其他组学层面的整合分析，为发育中的中枢神经系统表观基因组图谱研究提供了全新视角。本研究对小鼠胚胎干细胞（mouse ESC）与NIH-3T3细胞系开展了批量及单细胞G4 CUT&Tag实验（设置两个生物学重复），同时对小鼠脑组织实施了单细胞G4 CUT&Tag实验。