Consequences of myosin-1G dimerization on its regulated interaction with actin

肌球蛋白-1G 二聚化对其与肌动蛋白相互作用调节的影响

• 批准号: 9756194
• 负责人: Sarah Heissler
• 金额: $ 24.33万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9756194
• 关键词: Actin-Binding Protein; Actins; Activities of Daily Living; Actomyosin; Actomyosin Adenosinetriphosphatase; Adhesions; Advisory Committees; Amino Acids; Award; Binding; Binding Sites; Biochemical; Bioinformatics; Biological; Biological Assay; Biophysics; Bundling; CRISPR/Cas technology; Calmodulin; Calmodulin-Binding Proteins; Cell membrane; Cell physiology; Cells; Cellular biology; Chemicals; Closure by clamp; Communication; Complement; Cytoskeleton; Data; Development; Development Plans; Dimerization; Disease; Educational process of instructing; Educational workshop; Equilibrium; Extramural Activities; Funding; Goals; Grant; Health; Heart Diseases; Hematological Disease; Hematopoietic; Human; Human body; Image; Immune; In Vitro; Individual; Institution; K-Series Research Career Programs; K22 Award; Kinetics; Knowledge; Laboratories; Leadership; Link; Lobe; Lymphoid; Malignant Neoplasms; Measures; Mediating; Membrane; Mentors; Methods; Microfilaments; Microscopy; Modeling; Modernization; Molecular; Molecular Motors; Motor; Myosin ATPase; Myosin S-1; National Heart, Lung, and Blood Institute; Phase; Physiological; Postdoctoral Fellow; Preparation; Probability; Process; Proline; Property; Proteins; Research; Research Personnel; Research Training; Resolution; Site; Structure; System; T-Cell Activation; T-Lymphocyte; Techniques; Testing; TimeLine; Training; Training Activity; Tropomyosin; United States National Institutes of Health; Vocational Guidance; Work; Writing; base; biophysical techniques; career; career development; cell motility; crosslink; design; dimer; experience; genome editing; human disease; immunological synapse; immunological synapse formation; innovation; insight; knock-down; knockout gene; member; monomer; novel; overexpression; programs; reconstitution; research and development; response; single molecule; skills; spatiotemporal; student participation; success; virtual

 DESCRIPTION (provided by applicant): Project Summary/Abstract The actin cytoskeleton is a large-scale network that fills all cells of the human body. Through its dynamic interaction with myosin motors is it involved in virtually any cellular process and aberrant function of the actomyosin interaction is linked to numerous human diseases including cancers and diseases of the heart and blood. The Research Strategy of the proposed work is to determine the molecular details by which the virtually uncharacterized human hematopoietic myosin-1G undergoes a calmodulin-mediated dimerization to crosslink and bundle actin (Aim 1), to attribute kinetic parameters and actin binding properties to myosin-1G monomers and dimers (Aim 2), and to describe the physiological significance of the myosin-1G monomer-dimer equilibrium at the immunological synapse of lymphoid T-cells (Aim 3). Completion of the proposed Research Strategy will produce critical insights into a novel mode of myosin dimerization that causes the formation of emerging higher-order actin structures that will fundamentally advance our understanding of the interaction between myosin and actin at the molecular level. The candidate Dr. Heissler is currently a postdoctoral fellow at the NIH/NHLBI in the laboratory of Dr. James Sellers. Through this K22 Career Development Award proposal, the candidate seeks to systematically acquire additional mentored research training and career development training at the NIH/NHLBI through a detailed Career Development Plan designed to complement her current skill set. With the continued support of members of her Advisory Committee, the K22 Career Development Award will establish a training framework to initiate the research program in preparation for the candidate's independent career. Central part of the intramural phase of the K22 award will be the candidate's Advisory Committee that will evaluate her progress on the proposed research and career development training as outlined in the detailed Career Development Plan. The scientific training will support the proposed Specific Aims through a combination of specialized course work and hands-on training to complete the proposed innovative Research Strategy. New scientific skills that will be acquired include general training in cell biology techniques to study dynamic aspects of the actomyosin cytoskeleton including training in the in vitro activation of T-cells and the use of the modern genome-editing technique CRISPR for gene knockout. This specialized training in cell biology combined with training in super-resolution microscopy techniques to image single molecules in vitro and in cells with exceptional spatiotemporal resolution will systematically complement the candidate's current interdisciplinary skill set in biochemical and biophysical techniques and allow her to bridge the gap between in vitro single- molecule studies in reconstituted systems of purified proteins and live cells. Importantly, the techniques and approaches developed during the funding period of the award will not only advance our understanding of the interaction between myosin-1G and actin but also allow for the successful completion of the proposed Research Strategy. This will establish the basis of the candidate's first NIH R01 and additional independent funding applications upon her transition to independence. Besides the scientific training, the candidate will experience extensive career and professional training in the intramural phase of this award to master academic challenges in the extramural phase of the award. Training activities such as mentoring and teaching will be complemented with training in management and leadership in form of seminars and workshops. The professional career development training also involves mentoring of a post-baccalaureate student, participation in grant-writing workshops, development of communication skills, career counseling and assessment coaching to prepare the candidate at best for her transition to independence and her long-term goal of becoming a successful independent investigator at a prestigious academic institution.

 描述（由申请人提供）：项目摘要/摘要肌动蛋白细胞骨架是一个大规模的网络，充满了人体的所有细胞。通过其与肌球蛋白马达的动态相互作用，它参与几乎任何细胞过程，并且肌动球蛋白相互作用的异常功能与许多人类疾病有关，包括癌症以及心脏和血液疾病。拟议工作的研究策略是确定几乎未表征的人造血肌球蛋白-1G经历钙调蛋白介导的二聚化以交联和捆绑肌动蛋白的分子细节（目的1），将动力学参数和肌动蛋白结合特性归因于肌球蛋白-1G单体和二聚体（目的2），并描述肌球蛋白-1G单体-二聚体平衡在淋巴T细胞免疫突触中的生理意义（目的3）。完成拟议的研究策略将产生关键的见解到一个新的模式的肌球蛋白二聚化，导致形成新兴的高阶肌动蛋白结构，这将从根本上推进我们的理解肌球蛋白和肌动蛋白之间的相互作用在分子水平上。候选人Heissler博士目前是James Sellers博士实验室的NIH/NHLBI博士后研究员。通过这个K22职业发展奖提案，候选人寻求系统地获得额外的指导研究培训和职业发展培训在NIH/NHLBI通过详细的职业发展计划，旨在补充她目前的技能。在她的咨询委员会成员的持续支持下，K22职业发展奖将建立一个培训框架，启动研究计划，为候选人的独立职业生涯做准备。K22奖校内阶段的核心部分将是候选人的咨询委员会，该委员会将评估她在详细的职业发展计划中概述的拟议研究和职业发展培训方面的进展。科学培训将通过专业课程工作和实践培训相结合来支持拟议的具体目标，以完成拟议的创新研究战略。将获得的新的科学技能包括细胞生物学技术的一般培训，以研究肌动球蛋白细胞骨架的动态方面，包括T细胞体外激活的培训和使用现代基因组编辑技术CRISPR进行基因敲除。这种细胞生物学的专门培训与超分辨率显微镜技术的培训相结合，以在体外和细胞中以特殊的时空分辨率成像单分子，将系统地补充候选人目前在生物化学和生物物理技术方面的跨学科技能，并使她能够弥合纯化蛋白质和活细胞重建系统中体外单分子研究之间的差距。重要的是，在该奖项的资助期间开发的技术和方法不仅将推进我们对肌球蛋白-1G和肌动蛋白之间相互作用的理解，而且还允许成功完成拟议的研究策略。这将为候选人的第一个NIH R 01和她过渡到独立后的额外独立资金申请奠定基础。除了科学培训，候选人将在该奖项的校内阶段经历广泛的职业和专业培训，以掌握该奖项的校外阶段的学术挑战。除了辅导和教学等培训活动外，还将举办研讨会和讲习班，进行管理和领导能力培训。专业职业发展培训还包括指导一名学士后学生，参加赠款撰写讲习班，发展沟通技能，职业咨询和评估辅导，以使候选人最好为她向独立过渡以及她成为一名成功的独立调查员的长期目标做好准备。