Investigating the Role of the Helicase, IGHMBP2, and the RIG-I-Like Receptor Pathway in Spinal Muscular Atrophy with Respiratory Distress Type 1 and Neuromuscular Development (SMARD1)

项目摘要 免疫球蛋白hmu结合蛋白2基因（IGHMBP 2）中的有害隐性突变产生了一种新的免疫球蛋白。 运动神经元疾病（MND）的范围从不太严重的年轻成人发作的运动和感觉 神经性疾病，腓骨肌萎缩症2S型（CMT 2S），严重的，往往是致命的年轻人 脊髓性肌萎缩伴呼吸窘迫1型（SMARD 1）。IGHMBP 2突变 也与婴儿猝死综合症（SIDS）有关。由于这些疾病的罕见性， 许多研究已经投入到可能的治疗方法中，更不用说IGHMBP 2在引起糖尿病中的作用了。 表型IGHMBP 2被认为编码具有低持续合成能力的DNA/RNA解旋酶。它无处不在 在整个身体中以低水平表达，其中性腺和神经组织具有最高表达。它 也被认为在翻译中起作用，因为它与核糖体有关。通过建立 几种小鼠模型代表了人类患者中发现的疾病谱，我们在以下方面取得了进展： 了解IGHMBP 2不仅在这些疾病中发挥作用，而且在神经肌肉疾病中发挥作用。 开发/维护。我们最近对我们的严重SMARD 1和CMT 2S进行了RNA测序， 模型，并发现免疫系统RIG-I样受体（RLR）途径在脊髓中高度上调， 这些老鼠的尾巴该途径与典型的病毒双链RNA的检测相关， 通过RNase L导致感染细胞的最终死亡。我们还对MND小鼠模型进行了研究 在核输出介体因子编码基因Nemf中存在有害的隐性突变。在这些小鼠中， 我们看到了类似的疾病严重程度谱。NEMF蛋白因其在核糖体质量中的作用而闻名 控制在我们对突变的Nemf小鼠脊髓的RNAseq分析中，我们看到了类似的 RLR途径。我们还培育了Nemf和Ighmbp 2突变杂合的小鼠。而 杂合表型通常不会单独见于每个突变基因，这些双杂合突变体 表现出严重的MND表型。这表明这些突变影响相同的途径。我假设 IGHMBP 2功能失调和/或IGHMBP 2的减少导致某些与IGHMBP 2相关的RNA产物。 神经发育/维持建立，触发RLR通路并导致运动神经死亡 神经元这项研究将确定RLR通路是否对糖尿病患者有反应性、改善性或有害性影响。 神经肌肉变性这项研究开辟了一个新的类别的MND与RLR途径。我 提出了一系列与RLR敲除以及MND突变相关的遗传杂交。我也 建议使用单核RNAseq和RNAscope，一种单细胞荧光原位杂交测定， Ighmbp 2和Nemf突变小鼠的脊髓，以确定RLR信号的来源。

PROJECT ABSTRACT Deleterious recessive mutations in the immunoglobulin h mu-binding protein 2 gene (IGHMBP2) create a spectrum of motor neuron diseases (MNDs) ranging from the less severe young adult onset motor and sensory neuropathic disease, Charcot-Marie-Tooth disease type 2S (CMT2S), to the severe and often fatal young childhood disease, Spinal Muscular Atrophy with Respiratory Distress type 1 (SMARD1). IGHMBP2 mutations have also been associated with Sudden Infant Death Syndrome (SIDS). Due to the rarity of these diseases, not much research has been invested into possible therapeutics, let alone the role IGHMBP2 plays in causing the phenotype. IGHMBP2 is thought to encode a DNA/RNA helicase with low processivity. It is ubiquitously expressed at low levels throughout the body, with gonadal and nervous tissue having the highest expression. It is also believed to have a role in translation because of its association with ribosomes. Through the creation of several mouse models representing the spectrum of disease found in human patients, we have made strides in understanding that IGHMBP2 plays a role not only in these diseases, but in neuromuscular development/maintenance. We recently performed RNA sequencing on our severe SMARD1 and CMT2S models and found that the immune system RIG-I-like receptor (RLR) pathway is highly upregulated in the spinal cords of these mice. This pathway is associated with the detection of typically viral double-stranded RNA and causes eventual death of the infected cell via RNase L. We have also conducted studies on MND mouse models with deleterious recessive mutations in the nuclear export mediator factor-encoding gene Nemf. In these mice, we see a similar spectrum of disease severity. The NEMF protein is better known for its role in ribosome quality control. In our RNAseq analysis of the spinal cords of our mutated Nemf mice, we saw a similar upregulation of the RLR pathway. We also bred mice to be heterozygous for mutations in both Nemf and Ighmbp2. While a heterozygous phenotype is not typically seen for each mutated gene alone, these double heterozygous mutants showed a severe MND phenotype. This suggests that these mutations impact the same pathway. I hypothesize that dysfunctional IGHMBP2 and/or a decrease in IGHMBP2 causes certain RNA products associated with neural development/maintenance to build up, triggering the RLR pathway and causing death of the motor neurons. This study will determine if the RLR pathway has a reactive, ameliorative, or detrimental effect on neuromuscular degeneration. This study opens a novel category of MNDs associated with the RLR pathway. I propose a series of genetic crosses associated with knock-outs of RLR in addition to MND mutations. I also propose using single nuclear RNAseq and RNAscope, a single cell fluorescent in-situ hybridization assay, on the spinal cords of Ighmbp2 and Nemf mutant mice to determine the origin of the RLR signal.