Single cell ATAC-seq analysis of H3.3K4M WT and Hom brain samples (10X Genomics Multiome)

Methylation of lysine4 on Histone H3 (H3K4) is require for promoter and enhancer activation at gene loci and presents distinct methylation patterns over developmental gene clusters. Disrupting in the writer, eraser and reader of H3K4 methylation is associated with developmental syndromes. However, less studies focus on the role of H3K4 methylation in development of interneuron and hypothalamus. Here, we explore the role of H3K4 methylation in the medial ganglionic eminence (MGE) derive interneuron and hypothalamus by directly mutating the lysine 4 on H3.3 to the methionine in mice. Overall design: The Nkx2.1-Cre;H3.3K4M;Sun1-sfGFP mice were generated to harvest all Nkx2.1-lineage GFP+ nuclei via flow cytometry. Single nuclei from embryonic and adult samples from each genotype were prepared using 10X Genomics Multiome pipeline. Samples: E13.5 MGE and hypothalamus as well as P60 cortex and hypothalamus of H3.3K4M WT and Hom. In addition, one replicate for male and female for each condition.

组蛋白H3第4位赖氨酸的甲基化（H3K4 methylation）对于基因位点的启动子与增强子激活是必需的，且在发育基因簇上呈现独特的甲基化模式。H3K4甲基化的写入酶、擦除酶与读取蛋白功能异常，均与发育综合征密切相关。然而，目前鲜有研究聚焦于H3K4甲基化在中间神经元与下丘脑发育中的作用。本研究通过在小鼠体内直接将H3.3的第4位赖氨酸突变为甲硫氨酸，探究H3K4甲基化在内侧神经节隆起（medial ganglionic eminence, MGE）来源的中间神经元及下丘脑中的功能。实验设计概述：我们构建了Nkx2.1-Cre;H3.3K4M;Sun1-sfGFP小鼠，通过流式细胞术分选获取所有Nkx2.1谱系的GFP阳性细胞核。采用10X Genomics多组学流程，制备各基因型胚胎及成年样本的单细胞核悬液。实验样本涵盖：E13.5胎龄的内侧神经节隆起与下丘脑组织，以及P60日龄H3.3K4M野生型与纯合突变型小鼠的皮层与下丘脑组织。此外，每种实验条件均设置雌雄个体各1个生物学重复。