Department of Biomedical Engineering, University of Virginia, Charlottesville, Virginia.
Department of Microbiology, Immunology & Cancer Biology, University of Virginia, Charlottesville, Virginia.
Cancer Res. 2021 Apr 1;81(7):1868-1882. doi: 10.1158/0008-5472.CAN-20-1037. Epub 2021 Feb 2.
Cancer evolves from premalignant clones that adopt unusual cell states to achieve transformation. We previously pinpointed the oligodendrocyte precursor cell (OPC) as a cell of origin for glioma, but the early changes of mutant OPCs during premalignancy remained unknown. Using mice engineered for inducible - loss in OPCs, we acutely isolated labeled mutant OPCs by laser-capture microdissection, determined global gene-expression changes by bulk RNA sequencing, and compared with cell-state fluctuations at the single-cell level by stochastic profiling, which uses RNA-sequencing measurements from random pools of 10 mutant cells. At 12 days after - deletion, bulk differences were mostly limited to mitotic hallmarks and genes for ribosome biosynthesis, and stochastic profiling revealed a spectrum of stem-progenitor (), proneural, and mesenchymal states as potential starting points for gliomagenesis. At 90 days, bulk sequencing detected few differentially expressed transcripts, whereas stochastic profiling revealed cell states for neurons and mural cells that do not give rise to glial tumors, suggesting cellular dead-ends for gliomagenesis. Importantly, mutant OPCs that strongly expressed key effectors of nonsense-mediated decay () and homology-dependent DNA repair () were identified along with DNA-damage markers, suggesting transcription-associated replication stress. Analysis of 10-cell transcriptomes at 90 days identified a locus of elevated gene expression containing an additional repair endonuclease () and , a Ras-Raf antagonist and possible counterbalance to loss, which was microdeleted or downregulated in gliomas at 150 days. These hidden cell-state variations uncover replication stress as a potential bottleneck that must be resolved for glioma initiation. SIGNIFICANCE: Profiling premalignant cell states in a mouse model of glioma uncovers regulatory heterogeneity in glioma cells-of-origin and defines a state of replication stress that precedes tumor initiation..
癌症是从获得异常细胞状态以实现转化的癌前克隆演变而来的。我们之前已经确定少突胶质前体细胞 (OPC) 是神经胶质瘤的起源细胞，但在癌前状态下突变 OPC 的早期变化仍然未知。我们使用经过基因工程改造的可诱导 OPC 缺失的小鼠，通过激光捕获显微切割术急性分离标记的突变 OPC，通过批量 RNA 测序确定全局基因表达变化，并通过随机分组 10 个突变细胞的 RNA 测序测量值进行单细胞水平的细胞状态波动的随机分析进行比较。在 - 删除后的 12 天，批量差异主要局限于有丝分裂特征和核糖体生物合成基因，而随机分析揭示了一系列干细胞-祖细胞 (), 神经前体细胞和间充质状态，作为神经胶质瘤发生的潜在起点。在 90 天，批量测序检测到很少有差异表达的转录本，而随机分析则揭示了神经元和壁细胞的细胞状态，这些细胞状态不会导致神经胶质瘤，这表明神经胶质瘤发生的细胞死胡同。重要的是，还鉴定了强烈表达无意义介导的衰变 (NMD) 和同源依赖性 DNA 修复 () 的关键效应子的突变 OPC，以及 DNA 损伤标志物，提示转录相关的复制应激。在 90 天时对 10 个细胞转录组的分析确定了一个基因表达升高的基因座，其中包含一个额外的修复内切酶 () 和 ，它是 Ras-Raf 拮抗剂，可能与 缺失形成拮抗，在 150 天时，它在神经胶质瘤中发生微缺失或下调。这些隐藏的细胞状态变化揭示了复制应激作为启动神经胶质瘤所必需解决的潜在瓶颈。意义：在神经胶质瘤的小鼠模型中对癌前细胞状态进行分析，揭示了神经胶质瘤起源细胞的调控异质性，并定义了肿瘤起始前的复制应激状态。
