Cell and mechanobiology of Asymmetric Cell Division

不对称细胞分裂的细胞和力学生物学

• 批准号: 10550034
• 负责人: Clemens C Cabernard
• 金额: $ 37.77万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2027-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10550034
• 关键词: Acute; Affect; Behavior; Cell Size; Cell division; Cells; Central Nervous System; Centrosome; Chromatin; Cues; Cytoskeleton; Defect; Development; Drosophila genus; Generations; Immunohistochemistry; Interphase; Kinesin; Maintenance; Malignant Neoplasms; Medical; Microtubule-Organizing Center; Mitosis; Molecular; Myosin Type II; Neurodevelopmental Disorder; Organ; Organelles; Organism; Pathway interactions; Process; Proteins; RNA; Reproducibility; Research; Siblings; Sister; Sister Chromatid; Stereotyping; System; Tissues; Transcriptional Regulation; fly; in vivo; in vivo imaging; innovation; interest; live cell imaging; mechanical signal; nanobodies; nerve stem cell; neuroblast; non-muscle myosin; novel; optogenetics; programs; protein kinase N; public health relevance; rho GTP-Binding Proteins; segregation; spatiotemporal; stem cells; superresolution microscopy; tool; transcriptome sequencing; tumor

PROJECT SUMMARY Generating cells with different fates, functions and behaviors is critically important for the development and maintenance of tissues, organs, and multicellular organisms. Cellular diversity can be generated through Asymmetric Cell Division (ACD). Stem cells utilize ACD to create differentiating sibling cells while maintaining the stem cell in the process. In addition to the asymmetric partitioning of proteins or RNAs, other mechanisms such as mechanical cues, sibling cell size asymmetry or organelle asymmetry could potentially also contribute to binary cell fate decisions. Here, I propose to use asymmetrically dividing Drosophila neuroblasts, the neural stem cells of the developing fly central nervous system, to investigate the cell and mechanobiology of ACD in vivo. Recently, we discovered that Non-muscle Myosin II-dependent cortical flows, induced through both polarity- and spindle-dependent cues, are implicated in the generation of sibling cell size asymmetry. I will investigate how cortical flows are induced and modulated with spatiotemporal precision to achieve reproducible sibling cell size asymmetry. Our recent discovery of Protein Kinase N (PKN), and the Rho GTPase pathway as inducers of cortical flows will provide molecular entry points. I will also investigate how cell size asymmetry contributes to cell fate decisions, using RNA sequencing, immunohistochemistry, and long-term live cell imaging in vivo. A second project encompassed in this research direction is aimed at investigating the molecular mechanisms and function of molecular centrosome asymmetry, which is manifested in biased microtubule organizing center (MTOC) activity in interphase. We identified new proteins and mechanisms, such as Kinesins, Pp4 and dynamic centriolar protein localization in mitosis, regulating centrosome asymmetry. Centrosome segregation is highly stereotypic in stem cells, but whether and how centrosome asymmetry affects cell fate decisions, remains to be resolved. We will use fly neural stem cells to investigate the mechanisms and functions of centrosome asymmetry during ACD. I am particularly interested in investigating whether centrosome asymmetry provides a mechanism for biased cell fate determinant segregation, either via asymmetric RNA or sister chromatid segregation. I will also investigate whether biased MTOC activity impacts transcriptional regulation via chromatin organization. This research program will benefit from several novel and innovative tools, consisting of live cell imaging, superresolution microscopy, RNA sequencing and acute protein mislocalization and perturbation systems (nanobody, optogenetics), which my lab implemented to probe cytoskeletal dynamics with high spatial and/or temporal precision in vivo. ACD is an evolutionary conserved mechanism and the proposed research program is medically significant because defects in ACD can cause neurodevelopmental disorders or cancer.

项目概要 生成具有不同命运、功能和行为的细胞对于发育和发育至关重要。 组织、器官和多细胞生物体的维护。细胞多样性可以通过以下方式产生 不对称细胞分裂（ACD）。干细胞利用 ACD 来产生分化的兄弟细胞，同时 在此过程中维持干细胞。除了蛋白质或RNA的不对称分配外，其他 机械线索、兄弟细胞大小不对称或细胞器不对称等机制可能 也可能有助于二元细胞命运的决定。 在这里，我建议使用不对称分裂的果蝇神经母细胞，即果蝇的神经干细胞 开发果蝇中枢神经系统，研究体内 ACD 的细胞和力学生物学。最近， 我们发现非肌肉肌球蛋白 II 依赖性皮质流通过极性和 纺锤体依赖性线索与兄弟细胞大小不对称的产生有关。我将调查如何 以时空精度诱导和调节皮质流，以实现可重复的兄弟细胞 尺寸不对称。我们最近发现蛋白激酶 N (PKN) 和 Rho GTPase 通路作为诱导剂 皮质流将提供分子入口点。我还将研究细胞大小不对称如何影响 利用 RNA 测序、免疫组织化学和体内长期活细胞成像来决定细胞命运。 该研究方向的第二个项目旨在研究分子 分子中心体不对称的机制和功能，表现为偏向微管 组织中心 (MTOC) 间期活动。我们发现了新的蛋白质和机制，例如 有丝分裂中的驱动蛋白、Pp4 和动态中心粒蛋白定位，调节中心体不对称性。 中心体分离在干细胞中是高度刻板的，但是中心体是否以及如何分离 不对称性影响细胞命运决定，仍有待解决。我们将使用果蝇神经干细胞 研究 ACD 过程中中心体不对称的机制和功能。我特别感兴趣 研究中心体不对称是否提供了偏向细胞命运决定因素的机制 通过不对称 RNA 或姐妹染色单体分离。我也会调查是否有偏见 MTOC 活性通过染色质组织影响转录调控。 该研究计划将受益于多种新颖和创新的工具，包括活细胞成像、 超分辨率显微镜、RNA 测序和急性蛋白质错误定位和扰动系统 （纳米抗体，光遗传学），我的实验室用它来探测高空间和/或高空间和/或的细胞骨架动力学 体内时间精度。 ACD 是一种进化保守机制，拟议的研究计划在医学上 意义重大，因为 ACD 缺陷可能导致神经发育障碍或癌症。