A primed immune transcriptional program is activated in oligodendroglia in multiple sclerosis [Cut&Run IFNgvsCtr]

Multiple sclerosis (MS) is a disease characterized by a targeted immune attack on myelin in the central nervous system (CNS). We have previously shown that oligodendrocytes (OLs), myelin producing cells in the CNS, and their precursors (OPCs), acquire disease-specific transcriptional states in MS. To understand how these alternative transcriptional programs are activated in disease, we performed single-cell assay for transposase accessible chromatin using sequencing (scATAC-seq) on the OL lineage in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. We identified regulatory regions with increased accessibility in oligodendroglia (OLG) in EAE, some of which in the proximity of immune genes. A similar remodeling of chromatin accessibility was observed upon treatment of postnatal OPCs with interferon-gamma (IFNg), but not with dexamethasone. These changes in accessibility were not exclusive to distal enhancers, but also occurred at promoter regions, suggesting a role for promoters in mediating cell-state transitions. Notably, we found that a subset of immune genes already exhibited chromatin accessibility in OPCs ex vivo and in vivo, suggesting a primed chromatin state in OLG compatible with rapid transitions to an immune-competent state. Several primed genes presented bivalency of H3K4me3 and H3K27me3 at promoters in OPCs, with loss of H3K27me3 upon IFNg treatment. Inhibition of JMJD3/KDM6B, mediating removal of H3K27me3, led to the inability to activate these genes upon IFNg treatment. Importantly, OLGs from the adult human brain showed chromatin accessibility at immune gene loci, particularly at MHC-I pathway genes. A subset of single-nucleotide polymorphisms (SNPs) associated with MS susceptibility overlapped with these primed regulatory regions in OLG from both mouse and human CNS. Our data suggest that susceptibility for MS may involve activation of immune gene programs in OLG. These programs are under tight control at the chromatin level in OLG and may therefore constitute novel targets for immunological-based therapies for MS. We have performed bulk CUT&RUN for H3K27ac, H3K27me3, H3K4me3 and Ctcf from FACS sorted Sox10GFP+ oligodendrocyte precursor cells treated with IFN-gamma fro 48 hours.

多发性硬化症（multiple sclerosis, MS）是一种以中枢神经系统（central nervous system, CNS）内髓鞘被靶向免疫攻击为特征的疾病。我们此前已证实，中枢神经系统内的髓鞘生成细胞——少突胶质细胞（oligodendrocytes, OLs）及其前体细胞（oligodendrocyte precursor cells, OPCs），在多发性硬化症中会获得疾病特异性转录状态。为阐明此类替代性转录程序在疾病中的激活机制，我们针对多发性硬化症的实验性自身免疫性脑脊髓炎（experimental autoimmune encephalomyelitis, EAE）小鼠模型中的少突胶质细胞谱系，开展了单细胞转座酶可及性测序（single-cell assay for transposase accessible chromatin using sequencing, scATAC-seq）实验。我们在EAE模型的少突胶质细胞（oligodendroglia, OLG）中鉴定出染色质可及性升高的调控区域，其中部分区域紧邻免疫基因。用γ干扰素（interferon-gamma, IFNγ）处理产后OPCs后，可观察到类似的染色质可及性重塑，而地塞米松处理则无此效应。此类染色质可及性改变不仅局限于远端增强子，也发生在启动子区域，提示启动子在介导细胞状态转换中发挥作用。值得注意的是，我们发现部分免疫基因在体外和体内的OPCs中已呈现染色质可及性，这提示OLG中存在预启动染色质状态，可快速转换为免疫活性状态。部分预启动基因在OPCs的启动子区域呈现H3K4me3与H3K27me3的二价修饰状态，经IFNγ处理后会丢失H3K27me3修饰。介导H3K27me3去除的JMJD3/KDM6B抑制剂处理，会使细胞无法在IFNγ刺激下激活此类基因。重要的是，成人脑组织中的OLGs在免疫基因位点，尤其是主要组织相容性复合体I型（MHC-I）通路基因位点，呈现染色质可及性。部分与多发性硬化症易感性相关的单核苷酸多态性（single-nucleotide polymorphisms, SNPs），与小鼠和人类中枢神经系统OLGs中的此类预启动调控区域存在重叠。我们的研究数据表明，多发性硬化症的易感性可能与OLGs中免疫基因程序的激活有关。此类程序在OLGs中受到染色质层面的严格调控，因此或可成为多发性硬化症免疫疗法的新型靶点。我们对经IFNγ处理48小时的荧光激活细胞分选（fluorescence-activated cell sorting, FACS）得到的Sox10GFP阳性少突胶质前体细胞，开展了针对H3K27ac、H3K27me3、H3K4me3及Ctcf的批量CUT&RUN实验。