Mutations in X-linked methyl-CpG-binding protein 2 (MECP2) cause Rett syndrome (RTT). To identify functional pathways that could inform therapeutic entry points, we carried out a genetic screen for secondary mutations that improved phenotypes in Mecp2/Y mice after mutagenesis with N-ethyl-N-nitrosourea (ENU). Here, we report the isolation of 106 founder animals that show suppression of Mecp2-null traits from screening 3177 Mecp2/Y genomes. Whole-exome sequencing, genetic crosses, and association analysis identified 22 candidate genes. Additional lesions in these candidate genes or pathway components associate variant alleles with phenotypic improvement in 30 lines. A network analysis shows that 63% of the genes cluster into the functional categories of transcriptional repression, chromatin modification, or DNA repair, delineating a pathway relationship with MECP2. Many mutations lie in genes that modulate synaptic signaling or lipid homeostasis. Mutations in genes that function in the DNA damage response (DDR) also improve phenotypes in Mecp2/Y mice. Association analysis was successful in resolving combinatorial effects of multiple loci. One line, which carries a suppressor mutation in a gene required for cholesterol synthesis, Sqle, carries a second mutation in retinoblastoma binding protein 8, endonuclease (Rbbp8, also known as CtIP), which regulates a DDR choice in double-stranded break (DSB) repair. Cells from Mecp2/Y mice have increased DSBs, so this finding suggests that the balance between homology-directed repair and nonhomologous end joining is important for neuronal cells. In this and other lines, two suppressor mutations confer greater improvement than one alone, suggesting that combination therapies could be effective in RTT.

中文翻译：

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Mecp2 缺失小鼠的抑制突变暗示了 Rett 综合征病理学中的 DNA 损伤反应。


X 连锁甲基 CpG 结合蛋白 2 (MECP2) 的突变会导致 Rett 综合征 (RTT)。为了确定可以为治疗切入点提供信息的功能途径，我们对二次突变进行了遗传筛选，这些突变在用 N-乙基-N-亚硝基脲 (ENU) 诱变后改善了 Mecp2/Y 小鼠的表型。在此，我们报告了通过筛选 3177 个 Mecp2/Y 基因组分离出的 106 只始祖动物，这些动物显示出 Mecp2-null 性状受到抑制。全外显子组测序、遗传杂交和关联分析确定了 22 个候选基因。这些候选基因或途径成分中的其他损伤将变异等位基因与 30 个品系的表型改善相关联。网络分析显示，63% 的基因聚类为转录抑制、染色质修饰或 DNA 修复等功能类别，描绘了与 MECP2 的通路关系。许多突变存在于调节突触信号或脂质稳态的基因中。在 DNA 损伤反应 (DDR) 中起作用的基因突变也改善了 Mecp2/Y 小鼠的表型。关联分析成功解决了多个基因座的组合效应。其中一个品系携带胆固醇合成所需基因 Sqle 的抑制突变，同时携带视网膜母细胞瘤结合蛋白 8 核酸内切酶（Rbbp8，也称为 CtIP）的第二个突变，该突变调节双链断裂 (DSB) 中的 DDR 选择） 维修。 Mecp2/Y 小鼠的细胞 DSB 增加，因此这一发现表明同源定向修复和非同源末端连接之间的平衡对于神经元细胞很重要。在这一品系和其他品系中，两种抑制突变比单独一种抑制突变带来更大的改善，这表明联合疗法可能对 RTT 有效。