Mapping Brainstem Motor Neuron Subtypes and Genetic Pathways Involved in their Differential Susceptibility to Disease

项目摘要/摘要 这份NIH K08提案描述了一项为期五年的学术职业发展培训计划 神经病理学和发育神经遗传学。马修·罗斯博士已完成临床实习 布里格姆妇女医院(BWH)和波士顿的解剖病理学和神经病理学研究 哈佛医学院(HMS)的儿童医院(BCH)，并将着手一项研究计划 在神经学疾病导向研究方面为独立的学术生涯进行培训。 在这个培训项目中，Rose博士将获得深入研究眼运动神经元(OMN)的经验 发展和轴突靶向，解释单细胞RNA测序，基因调控分析 网络、组织清理和先进的成像方法以及基因编辑技术。他的导师，斯图尔特博士。 伊丽莎白·恩格尔(BCH)神经学教授、HHMI调查员，脑干OMN领域的领军人物 发育和先天性颅神经紊乱(CCDD)，涉及这些运动神经元。她 实验室在人类和小鼠遗传学、神经解剖学、大数据分析、 分子生物学、成像和细胞培养技术。她在指导其他人方面有既定的记录 受训人员将在生物医学研究领域取得成功。除了他的导师，他还组建了一个小组 在这些学科领域拥有互补专业知识的合作者，以及由资深教师组成的咨询委员会 在指导内科科学家向独立过渡方面拥有丰富的经验。他会的 在这项培训的基础上增加教学内容，并在全国会议上介绍工作。 提出的研究计划的主要科学目标是研究正常和异常的OMN规范 以及轴突生长和对健康和疾病的指导。罗斯博士提供了一张遗传基因的初步图谱 脑干运动神经元的差异可能成为进一步研究所有运动神经元的基础 疾病。这些初步数据将作为进一步研究OMN发展的基础 通过发现每个OMN亚群的标记，然后将这些标记应用于对OMN的研究 CCDD。这一建议的中心假设是OMN之间基因表达的差异 在不同的CCDD中，亚群可能倾向于某些亚型经历神经功能障碍。 单细胞rna-seq将在三个主要的动眼神经上进行：动眼神经、滑车神经和外展神经。 脑干核团，以发现每个亚群的标记。与ATAC-seq相结合， 将对OMN规范的转录调控网络进行调查。每个OMN的唯一标记 然后将使用亚群来研究正常的OMN轴突向不同肌肉靶点的投射，以及 用它们来剖析不同人群对刻板印象和病理性异常的贡献 在CCDD中观察到神经支配。这些研究对神经细胞的更广泛领域具有重要意义。 规范和分化，并对轴突的生长和引导。

PROJECT SUMMARY/ABSTRACT This NIH K08 proposal describes a five-year training program for career development in academic neuropathology and developmental neurogenetics. Dr. Matthew Rose has completed clinical residency in Anatomic Pathology and fellowship in Neuropathology at Brigham and Women’s Hospital (BWH) and Boston Children’s Hospital (BCH) at Harvard Medical School (HMS), and will embark on a research program designed to train for an independent academic career in neurologic disease-oriented research. In this training program, Dr. Rose will acquire in-depth experience in the study of ocular motor neuron (OMN) development and axon targeting, interpretation of single cell RNA-sequencing, analysis of gene regulatory networks, tissue clearing and advanced imaging approaches, and gene-editing technologies. His mentor, Dr. Elizabeth Engle (a Professor of Neurology and HHMI Investigator at BCH), is a leader in brainstem OMN development and congenital cranial dysinnervation disorders (CCDDs) that involve these motor neurons. Her laboratory has extensive experience in human and mouse genetics, neuroanatomy, large data analyses, molecular biology, imaging, and cell culture techniques. She has an established track record for mentoring other trainees to successful careers in biomedical investigation. In addition to his mentor, he has assembled a group of collaborators with complementary expertise in these disciplines, and an Advisory Committee of senior faculty with extensive experience in guiding physician-scientists through the transition to independence. He will supplement this training with didactics and presentation of work at national meetings. The primary scientific goal of the proposed research plan is to study normal and abnormal OMN specification and axonal growth and guidance in health and disease. Dr. Rose provides an initial map of the genetic differences among brainstem motor neurons that may serve as a foundation for further studies of all motor neuron diseases. These preliminary data will be used as a foundation to further study the development of the OMN subnuclei by discovering markers of each OMN subpopulation, and then applying those markers to the study of the CCDDs. The central hypothesis of this proposal is that the differences in gene expression among OMN subpopulations may predispose certain subtypes to undergo dysinnervation in different CCDDs. Single cell RNA-seq will be performed on the three primary OMNs: the oculomotor, trochlear, and abducens brainstem nuclei, in order to discover markers of each subpopulation. In combination with ATAC-seq, the transcriptional regulatory networks for OMN specification will be investigated. Unique markers of each OMN subpopulation will then be used to study normal OMN axon projections to distinct muscle targets, as well as using them to dissect the contributions of different populations to the stereotypic and pathologic aberrant innervations observed in the CCDDs. These studies have significance to the broader fields of neuronal specification and differentiation and to axonal growth and guidance.