CHARACTERIZATION OF MOB1 DYNAMICS IN LIVING CELLS

活细胞中 MOB1 动力学的表征

• 批准号: 7960229
• 负责人: Charles Bradley Shuster
• 金额: $ 12.87万
• 依托单位: NEW MEXICO STATE UNIVERSITY LAS CRUCES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7960229
• 关键词: Anaphase; Binding; Biomedical Research; CDC2 Protein Kinase; Cell division; Cells; Chimera organism; Chromosome Segregation; Chromosomes; Computer Retrieval of Information on Scientific Projects Database; Cytokinesis; Eukaryota; Family member; Foundations; Funding; Grant; Homologous Gene; Human; Institution; Kinetochores; LATS2 gene; Lead; Life; Maintenance; Mammalian Cell; Mitosis; Mitotic; Mitotic Checkpoint; Monitor; Names; New Mexico; Phosphotransferases; Play; Ploidies; Research; Research Personnel; Resources; Role; Signal Transduction; Sister Chromatid; Small Interfering RNA; Source; United States National Institutes of Health; Yeasts; cellular imaging; fungus; in vivo; mutant; novel; segregation

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Maintenance of chromosomal ploidy during cell division requires a precise coordination of chromosome segregation and cytoplasmic partitioning (cytokinesis) such that the contractile ring does not assemble before the onset of anaphase. Progression of dividing cells into anaphase into cytokinesis is dependent on the spindle assembly checkpoint, which monitors chromosome attachment to the spindle and regulates the onset of sister chromatid segregation and CDK1 inactivation (mitotic exit). In yeast, the mitotic checkpoint also regulates a signaling cascade termed the Septation Initiation Network (SIN) that regulates the initiation of cytokinesis. Few functional homologues of the SIN have been identified in higher eukaryotes, but a terminal component named Mob1 has been identified in all eukaryotes. Expression of GFP chimeras of the four human Mob1s reveal that Mob1 localizes to the spindle poles and kinetochores up until anaphase onset, and the central spindle and midbody during cytokinesis. Additionally, we have identified a novel interaction between Mob1A and LATS2 kinase, which may serve as the functional homolog of the yeast Dbf2/Sid2 kinase in mammalian cells. Using these preliminary studies as a foundation, this application seeks to evaluate the roles that Mob1 plays in coordinating mitosis and cytokinesis in mammalian cells. The lines of experimentation proposed in this application will characterize the localization dynamics of the four human Mob1 family members using co-localization studies and live cell imaging. Further, the role of Mob1 in the coordination of mitosis and cytokinesis will be assessed by specifically disrupting Mob1/Lats2 interactions in vivo by siRNA knockdown of Mob1 or by expression of Lats2-binding mutants of Mob1. These efforts should ultimately lead to a clearer understanding of how mitosis and cytokinesis is coordinated, and confirm whether the strategies employed by unicellular fungi to regulate septation are conserved in higher eukaryotes.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 在细胞分裂期间维持染色体倍性需要染色体分离和细胞质分配（胞质分裂）的精确协调，使得收缩环在后期开始之前不组装。分裂细胞进入后期到胞质分裂的进展依赖于纺锤体组装检查点，该检查点监测染色体附着到纺锤体并调节姐妹染色单体分离和CDK 1失活（有丝分裂退出）的开始。在酵母中，有丝分裂检查点还调节称为分隔起始网络（SIN）的信号级联，其调节胞质分裂的起始。在高等真核生物中，SIN的功能同源物很少，但在所有真核生物中都发现了一个名为Mob 1的末端组分。四个人Mob 1的GFP嵌合体的表达揭示Mob 1定位于纺锤体极和动粒，直到后期开始，以及在胞质分裂期间的中央纺锤体和中间体。此外，我们已经确定了Mob 1A和LATS 2激酶之间的一种新的相互作用，这可能是哺乳动物细胞中酵母Dbf 2/Sid 2激酶的功能同源物。利用这些初步研究作为基础，本申请旨在评估Mob 1在协调哺乳动物细胞有丝分裂和胞质分裂中发挥的作用。本申请中提出的实验路线将使用共定位研究和活细胞成像来表征四个人类Mob 1家族成员的定位动力学。此外，Mob 1在协调有丝分裂和胞质分裂中的作用将通过特异性破坏Mob 1/Lats 2相互作用来评估，所述相互作用通过siRNA敲低Mob 1或通过表达Mob 1的Lats 2结合突变体来体内进行。这些努力应最终导致有丝分裂和胞质分裂是如何协调的更清楚的理解，并确认是否单细胞真菌采用的策略，以调节分隔是保守的高等真核生物。