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(申请人提供)：项目摘要/摘要肌动蛋白细胞骨架是一个充满人体所有细胞的大规模网络。通过它与肌球蛋白马达的动态相互作用，它几乎参与了任何细胞过程，而肌动蛋白相互作用的异常功能与许多人类疾病有关，包括癌症和心脏和血液疾病。这项拟议工作的研究策略是确定几乎没有特征的人造血肌球蛋白-1G经历钙调蛋白介导的二聚化为交联和捆绑肌动蛋白的分子细节(目标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(目标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学意义(目标3)。拟议的研究策略的完成将对肌球蛋白二聚化的新模式产生关键的见解，这种模式导致新的高阶肌动蛋白结构的形成，这将从根本上促进我们在分子水平上理解肌球蛋白和肌动蛋白之间的相互作用。候选人海斯勒博士目前是美国国立卫生研究院/NHLBI詹姆斯·塞勒斯博士实验室的博士后研究员。通过这项K22职业发展奖提案，候选人寻求通过详细的职业发展计划系统地在NIH/NHLBI获得额外的指导研究培训和职业发展培训，以补充她目前的技能集。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，以启动研究计划，为候选人的独立职业生涯做准备。K22奖项内部阶段的中心部分将是候选人咨询委员会，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情况。科学培训将通过专业课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。将获得的新科学技能包括细胞生物学技术的一般培训，以研究肌动蛋白细胞骨架的动态方面，包括培训T细胞的体外激活，以及使用现代基因组编辑技术CRISPR进行基因敲除。这一专门的细胞生物学培训与超分辨率显微技术的培训相结合，在体外和在具有非凡时空分辨率的细胞中成像单分子，将系统地补充应聘者目前在生化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组系统中体外单分子研究之间的差距。重要的是，在该奖项的资助期内开发的技术和方法不仅将促进我们对肌球蛋白-1G和肌动蛋白之间相互作用的理解，而且还将使拟议的研究战略顺利完成。这将为候选人过渡到独立后的第一个NIH R01和其他独立资金申请奠定基础。除了科学培训外，候选人还将在该奖项的校内阶段经历广泛的职业和专业培训，以掌握该奖项校外阶段的学术挑战。指导和教学等培训活动将以研讨会和讲习班的形式辅之以管理和领导方面的培训。职业生涯发展培训还包括指导一名毕业后的学生，参加撰写补助金的研讨会，发展沟通技能，职业咨询和评估指导，为候选人过渡到独立和她成为一家著名学术机构成功的独立调查员的长期目标做好最好的准备。