SINGLE-CELL ANALYSIS OF SOMATIC MUTATION IN AGING ANO NEUROOEGENERATIVE DISEASE IN THE HUMAN BRAIN

人脑衰老和神经生成疾病中体细胞突变的单细胞分析

• 批准号: 10237914
• 负责人: Michael Anthony Lodato
• 金额: $ 24.58万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10237914
• 关键词: Adult; Affect; Age; Aging; Alzheimer' s Disease; Alzheimer' s disease pathology; Alzheimer' s disease patient; Antibiotics; Antioxidants; Authorship; Award; Base Pairing; Bioinformatics; Boston; Brain; Cardiovascular Diseases; Cells; Cerebral cortex; Characteristics; Cockayne Syndrome; Communities; DNA; DNA Damage; DNA Markers; DNA Sequence Alteration; DNA sequencing; Data; Development; Diabetes Mellitus; Disease; Doctor of Medicine; Doctor of Philosophy; Educational workshop; Elderly; Environment; Epigenetic Process; Equipment; Faculty; Fellowship; Genes; Genetic; Genetic Transcription; Genome; Genomics; Genotoxic Stress; Goals; Health; Human; Human Genome Project; Human body; Incidence; Individual; Inherited; Institutes; Institution; Laboratories; Lesion; Life; Malignant Neoplasms; Manuscripts; Massachusetts; Mediating; Mentors; Mitotic; Molecular Biology; Mutation; Mutation Analysis; Nerve Degeneration; Nervous System Physiology; Nervous system structure; Neurobiology; Neurodegenerative Disorders; Neurons; Nucleotides; Oxidative Stress; Paper; Pathogenicity; Pathologic Mutagenesis; Pathway interactions; Patients; Pattern; Pediatric Hospitals; Peer Review; Persons; Phase; Phenotype; Positioning Attribute; Postdoctoral Fellow; Production; Progeria; Proteins; Publications; Publishing; Reactive Oxygen Species; Reagent; Research; Research Personnel; Research Training; Risk Factors; Role; Sampling; Scanning Electron Microscopy; Science; Secure; Single Nucleotide Polymorphism; Somatic Mutation; Supervision; Technology; Testing; Tissue Culture Techniques; Tissues; Universities; Variant; Walking; Work; Xeroderma Pigmentosum; age group; age related; aging brain; base; body system; career; career development; cognitive function; experience; experimental study; genome integrity; genome sequencing; human disease; interest; loss of function; loss of function mutation; medical schools; meetings; novel; oxidative DNA damage; post-doctoral training; repaired; response; single cell analysis; single cell sequencing; skills; stem; success; supportive environment; tenure track; transdifferentiation; undergraduate student; virtual; whole genome

Project Summary/Abstract Candidate. I have a profound interest in understanding the genetic and epigenetic mechanisms that control cell state, and have built upon that interest during my research career. As an undergraduate researcher in the laboratory of Dr. Joanne Wiley at Hofstra University, I studied bacterial sporulation and antibiotic production using genetics and scanning electron microscopy, resulting in authorship on a peer-reviewed manuscript. As a Ph.D. candidate in the laboratory of Dr. Rudolf Jaenisch at the Massachusetts Institute of Technology, I worked on several of projects which resulted in authorship of peer-reviewed manuscripts, related to epigenetics, factor- mediated reprogramming, and transdifferentiation. In my main thesis work, I used genomics, genetics, molecular biology, and tissue culture techniques to study cell fate transitions in the development of the mammalian nervous system, resulting in a first-author publication (Lodato et al., PLoS Genetics, 2013, PMID 23437007). In the first part of my postdoctoral fellowship, in the lab of Dr. Christopher Walsh of Boston Children’s Hospital and Harvard Medical School, I built upon all these previous experiences, in particular in using genomics to understand principles of neurobiology, to study somatic mutations in the normal human brain using cutting-edge single-cell whole genome sequencing (WGS) technologies, resulting in a first-author manuscript (Lodato et al., Science, 2015, PMID 26430121). This paper was the first published work to compare the whole-genome rates and characteristics of somatic mutations across normal human individuals, and the expertise I gained developing that technology has prepared me thoroughly to now compare the rates and characteristics of somatic mutations between age groups and in human age-related diseases. The K99/R00 Pathway to Independence Award is the perfect mechanism to help propel me to my scientific and career goals. Under this award, I will 1) Execute a research plan to uncover the role of somatic mutation in aging and in age-related diseases 2) Gain scientific skills and refine my understanding of key concepts in the fields of aging and bioinformatics 3) Undertake career development activities under the supervision of my mentor, co-mentor, and other faculty advisors committed to my successful transition from postdoctoral fellow to independent investigator. Environment. The proposed Research and Training plans will take place in the laboratory of Christopher A. Walsh, M.D., Ph.D., within Boston Children’s Hospital (BCH) and Harvard Medical School (HMS). These institutions comprise a strong, well-established research community, and are committed to the success of the aims and goals proposed in this application. Dr. Walsh has a strong track record of training postdoctoral fellows, many of his former trainees now holding tenured or tenure-track faculty positions at academic institutions. This success stems from the rigorous and supportive environment in the lab, including frequent lab meetings and one-on-one interactions with Dr. Walsh. Scientifically, the lab is supplied with virtually all reagents and equipment needed for the proposed research, and as a part of the greater BCH and HMS community any additional equipment are available within walking distance. The BCH/HMS community is also an asset to the career development aspects of this proposal, providing easy access to potential collaborators and frequent seminars hosting internal and external speakers. As part of this community, and I have secured a co-mentor (Dr. Peter Park, HMS), and two additional faculty advisors (Drs. Bruce Yanker, HMS, and Emanuela Gussoni, BCH/HMS), all committed to my success. Finally, the BCH/HMS environment provides myriad formal career development seminars and workshops, which I will take part in to aid my transition to independence. Research. Advanced age is a major risk factor for human diseases in all organ systems, yet how the phenomenon of aging affects such a wide spectrum of tissues is unknown. All tissues rely on the integrity of the genome to function properly, and one attractive and long-standing hypothesis is that the gradual accumulation of DNA damage might be causal in aging. Proof of this notion has remained elusive in the brain, since standard DNA-sequencing experiments are ill-suited to detect somatic mutations which might mark only a few cells, or even a single cell, in a sample comprised of millions. I have pioneered the use of single-cell, whole-genome sequencing technology to compare rates, characteristics, and consequences of somatic mutations across human brains, and will use this technology as a part of this award to achieve three Aims: 1) Determine whether aging is associated with an accumulation of mutations in the human brain and define patterns of somatic mutation during aging, 2) Single-cell sequencing in progeroid diseases associated with DNA-damage response, and 3) Examination of the somatic mutation rate in Alzheimer’s disease. Thus, this proposal represents a comprehensive body of work that will elucidate the role of somatic mutation in during aging, in accelerated aging, and in age-associated disease in the human brain.

项目总结/摘要 候选人我对理解控制人类基因组的遗传和表观遗传机制 细胞状态，并在我的研究生涯中建立了这种兴趣。作为一名本科研究员， 在霍夫斯特拉大学乔安妮·威利博士的实验室里，我研究了细菌孢子形成和抗生素生产 使用遗传学和扫描电子显微镜，在同行评审的手稿上产生作者身份。作为 博士作为马萨诸塞州理工学院鲁道夫·耶尼施博士实验室的候选人， 在几个项目，导致作者的同行评议的手稿，有关表观遗传学，因素- 介导的重编程和转分化。在我的主要论文中，我使用了基因组学，遗传学， 分子生物学和组织培养技术来研究细胞命运的转变， 哺乳动物神经系统，导致第一作者出版物（Lodato等人，PLoS Genetics，2013，PMID 23437007）。在我博士后研究的第一部分，在波士顿的克里斯托弗沃尔什博士的实验室里， 儿童医院和哈佛医学院，我建立在所有这些以前的经验，特别是在 利用基因组学来理解神经生物学原理，研究正常人的体细胞突变， 大脑使用尖端的单细胞全基因组测序（WGS）技术，导致第一作者 手稿（Lodato等人，Science，2015，PMID 26430121）。这篇论文是第一篇发表的工作， 比较正常人类个体的全基因组率和体细胞突变的特征， 我在开发这项技术时所获得的专业知识使我做好了充分的准备， 以及年龄组之间和人类年龄相关疾病中体细胞突变的特征。 K99/R 00独立之路奖是一个完美的机制，帮助推动我实现我的目标。 科学和职业目标。在这个奖项下，我将1）执行一项研究计划，以揭示体细胞的作用， 2）获得科学技能，完善我对衰老和与年龄有关的疾病的关键的理解 3）在老龄化和生物信息学领域开展职业发展活动， 我的导师，共同导师和其他教师顾问的监督，致力于我的成功过渡， 从博士后到独立调查员 环境拟议的研究和培训计划将在Christopher A. 医学博士沃尔什，哲学博士、波士顿儿童医院（BCH）和哈佛医学院（HMS）。这些 各院校组成了一个强大的，完善的研究社区，并致力于成功的 本申请提出的目的和目标。沃尔什博士在培养博士后方面有着良好的记录 研究员，他的许多前学员现在持有终身或终身教职的学术职位 机构职能体系这一成功源于实验室中严格和支持性的环境，包括频繁的实验室 与沃尔什医生的会议和一对一的互动。从科学角度讲，实验室几乎提供了 拟议研究所需的试剂和设备，以及作为更大的BCH和HMS的一部分 社区任何额外的设备都可以在步行距离内。BCH/HMS社区也是 这是本提案职业发展方面的一项资产，可以方便地接触到潜在的合作者 并经常举办研讨会，邀请内部和外部发言人。作为这个社区的一部分，我已经获得了一个 共同导师（博士彼得公园，HMS），和两个额外的教师顾问（博士布鲁斯扬克，HMS和埃马努埃拉 Gussoni，BCH/HMS），都致力于我的成功。最后，BCH/HMS环境提供了大量的正式的 职业发展研讨会和讲习班，我将参加这些研讨会和讲习班，以帮助我向独立过渡。 Research.高龄是所有器官系统中人类疾病的主要危险因素， 老化现象影响如此广泛的组织是未知的。所有组织都依赖于 基因组正常运作，一个有吸引力的和长期存在的假设是， DNA损伤的积累可能是衰老的原因。这一概念的证据在大脑中仍然难以捉摸， 由于标准DNA测序实验不适合检测可能仅标记 几个细胞，甚至是一个细胞，在一个由数百万个细胞组成的样本中。我开创了单细胞的应用， 全基因组测序技术，以比较率，特点，和结果的体细胞 人类大脑中的突变，并将使用这项技术作为该奖项的一部分，以实现三个目标：1） 确定衰老是否与人类大脑中突变的积累有关，并定义 衰老过程中体细胞突变的模式，2）与衰老相关的早老性疾病中的单细胞测序 DNA损伤反应，和3）阿尔茨海默病中体细胞突变率的检查。因此，这 该提案代表了一个全面的工作机构，将阐明体细胞突变在过程中的作用， 老化，加速老化，以及与年龄相关的疾病在人类大脑中。

项目成果

Persistent DNA damage associated with ATM kinase deficiency promotes microglial dysfunction.

• DOI: 10.1093/nar/gkac104
• 发表时间: 2022-03-21
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Bourseguin J;Cheng W;Talbot E;Hardy L;Lai J;Jeffries AM;Lodato MA;Lee EA;Khoronenkova SV
• 通讯作者: Khoronenkova SV

Single-cell transcriptomic and genomic changes in the aging human brain.

衰老人脑中的单细胞转录组和基因组变化。

• DOI: 10.1101/2023.11.07.566050
• 发表时间: 2023
• 期刊: bioRxiv : the preprint server for biology
• 作者: Jeffries,AilsaM;Yu,Tianxiong;Ziegenfuss,JenniferS;Tolles,AllieK;Kim,Yerin;Weng,Zhiping;Lodato,MichaelA
• 通讯作者: Lodato,MichaelA