Studies of Hereditary Neurological Disease: Disease Mechanisms

最近，我们的研究集中在三种遗传性运动神经元病上：由于感觉神经毒素(SETX)突变引起的肌萎缩侧索硬化症(ALS4)，由于雄激素受体(AR)突变引起的脊髓和延髓肌萎缩(SBMA)，以及由于SMN蛋白缺失引起的常染色体隐性遗传性脊肌萎缩(SMA)。具体研究成果包括： (1)我们研究了由SETX已知突变引起的ALS4患者的临床和分子特征，并开发了评估其他SETX变异的工具。我们的研究涉及32例患者，包括31例SETX基因C.1166 T&gt；C突变(p.Leu389Ser)和1例C.1153 G&gt；A突变(p.Glu385Lys)。患者的临床特征包括神经学检查、血液检查、磁共振成像(MRI)和双能X射线吸收法(DEXA)。取成纤维细胞和运动神经元建立模型，研究其分子功能的变化。我们报告了ALS4的主要临床特征。实验室分析显示，Leu389Ser ALS4队列中血清肌酸激酶和肌酐发生变化。MRI显示双侧肢体肌肉脂肪分数增加，与病程相关。DEXA测量显示，下肢比上肢受到的影响更大。对SETX功能的细胞检测证实，与Leu389Ser突变一样，Glu385Lys变异导致R环减少，可能是由于功能的增加。我们确定了ALS4的临床实验室和放射学特征，因此应该监测它们的疾病进展。患者来源细胞中R-loop水平的分子特征为深入了解疾病病理和评估SETX基因候选突变的致病性提供了依据。 (2)SBMA是由AR蛋白中的多聚谷氨酰胺膨胀引起的。利用基因表达分析和芯片测序，我们绘制了基因工程患者干细胞来源运动神经元的转录变化图谱。我们发现SBMA的转录失调可以通过AR介导组蛋白修饰来发生。我们检测到组蛋白乙酰化减少，编码代偿代谢蛋白的基因表达减少，线粒体功能的底物可用性降低。此外，我们发现补充丙酮酸纠正了这一缺陷，并改善了线粒体功能和SBMA运动神经元活性。我们认为代谢基因的表观遗传失调是导致线粒体ATP产量减少的原因。我们的结果表明，表观遗传调控的改变与导致神经变性的线粒体新陈代谢之间存在分子联系。

Recently our research has focused on three hereditary motor neuron diseases: amyotrophic lateral sclerosis type 4 (ALS4) due to mutation in senataxin (SETX), spinal and bulbar muscular atrophy (SBMA) due to mutation in the androgen receptor (AR), and autosomal recessive spinal muscular atrophy (SMA) due to deficiency of the protein SMN. Specific research accomplishments include the following: (1) We characterized clinical and molecular features in patients with ALS4 caused by known mutation in SETX, and we developed tools for evaluating other SETX variants. Our study involved 32 patients, including 31 with mutation in SETX at c.1166 T>C (p.Leu389Ser) and 1 with mutation at c.1153 G>A (p.Glu385Lys). Clinical characterization of the patients included neurological examination, blood tests, magnetic resonance imaging (MRI), and dual-energy x-ray absorptiometry (DEXA). Fibroblasts and motor neurons were obtained to model the disease and characterize the molecular alteration in senataxin function. We reported key clinical features of ALS4. Laboratory analysis showed alteration of serum creatine kinase and creatinine in the Leu389Ser ALS4 cohort. MRI showed increased muscle fat fraction in the lower extremities, which correlated with disease duration. DEXA measurements showed that lower extremities are more affected than upper extremities. A cellular assay for SETX function confirmed that like the Leu389Ser mutation, the Glu385Lys variant leads to a decrease in R loops, likely from a gain of function. We identified clinical laboratory and radiological features of ALS4, and hence they should be monitored for disease progression. The molecular characterization of R-loop levels in patient-derived cells provides insight into the disease pathology and assays to evaluate the pathogenicity of candidate mutations in the SETX gene. (2) SBMA is caused by polyglutamine expansion in the AR protein. Using gene expression analysis and ChIP sequencing, we mapped transcriptional changes in genetically engineered patient stem cell-derived motor neurons. We found that transcriptional dysregulation in SBMA can occur through AR-mediated histone modification. We detected reduced histone acetylation, along with decreased expression of genes encoding compensatory metabolic proteins and reduced substrate availability for mitochondrial function. In addition, we found that pyruvate supplementation corrected this deficiency and improved mitochondrial function and SBMA motor neuron viability. We propose that epigenetic dysregulation of metabolic genes contributes to reduced mitochondrial ATP production. Our results show a molecular link between altered epigenetic regulation and mitochondrial metabolism that results in neurodegeneration.