IDENTIFICATION OF KINETOCHORE INTERACTING PROTEINS (KNL-1/KNL-3/KNL-2)

动粒相互作用蛋白的鉴定 (KNL-1/KNL-3/KNL-2)

• 批准号: 8171385
• 负责人: Arshad Desai
• 金额: $ 0.24万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8171385
• 关键词: Biological Assay; Caenorhabditis elegans; Cell division; Chromosome Segregation; Chromosomes; Collaborations; Computer Retrieval of Information on Scientific Projects Database; Congenital Abnormality; Consensus; Embryo; Funding; Grant; In Vitro; Institution; Kinetochores; Location; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Mitosis; Mitotic; Mitotic Checkpoint; N-terminal; Phosphorylation; Phosphotransferases; Physiological; Proteins; Proteomics; Radioactive; Regulation; Research; Research Institute; Research Personnel; Resources; Role; Sampling; Series; Signal Transduction; Site; Source; Testing; Time; United States National Institutes of Health; human PLK1 protein; melting; prevent

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. Polo-like kinase 1 (Plk1) is a central mitotic kinase implicated in the regulation of the mitotic checkpoint, a mechanism controlling chromosomes segregation and preventing loss or gain of whole chromosomes, which is a known cause of birth defects and a contributing factor in the genesis of cancer. The multiplicity of Plk1 functions for has made it difficult to deconvolve its function in cell division. A potential approach to circumventing this problem is to identify kinetochore-specific PLK-1 substrates. We identified a series of N-terminal ?MELT? repeats in a protein called KNL-1 (Kinetochore Null protein-1) that fit the consensus site for PLK-1 phosphorylation. KNL-1 is a protein located on the chromosomes necessary for the assembly of the kinetochore and chromosomes segregation. We are testing the hypothesis that KNL-1 is a PLK-1 substrate and investigate the role of this phosphoregulation in chromosome segregation and checkpoint signaling. As predicted by the location of the MELT repeats, the N-terminal fragment of KNL-1 (KNL-11-505) is an excellent PLK-1 substrate in vitro (radioactive kinase assays). To assess the function of PLK-1 phosphorylation of KNL-1, in collaboration with the Scripps Research Institute Center for Physiological Proteomics, we mapped PLK-1 target sites on KNL-1 by using a combination of in vitro kinase assays and analysis by mass spectrometry. 6 phosphosites were each identified multiple times in the 3 samples we submitted to analysis, indicating that they are major sites on KNL-1 for PLK-1 phosphorylation. Now we are investigating the role of these phosphorylations in the control of mitosis progression and chromosomes segregation in C.elegans embryos.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 polo样激酶1（PLK1）是一种涉及有丝分裂检查点的调节的中心有丝分裂激酶，一种控制染色体隔离的机制，可预防整个染色体的损失或增益，这是已知的出生缺陷的原因，并且是癌症创建中的造成生育缺陷的原因。 PLK1功能的多样性使得很难将其在细胞分裂中的功能解析。解决此问题的潜在方法是识别动力学特异性PLK-1底物。我们确定了一系列N端？熔体？在称为PLK-1磷酸化共有位点的称为KNL-1（Kinetochore Null蛋白1）的蛋白质中重复。 KNL-1是一种位于动物学和染色体隔离所需的染色体上的蛋白质。我们正在检验以下假设：KNL-1是PLK-1底物，并研究了该磷酸盐调节在染色体分离和检查点信号传导中的作用。正如熔体重复序列的位置所预测的那样，KNL-1（KNL-11-505）的N末端片段是体外的极好的PLK-1底物（放射性激酶测定）。为了评估KNL-1的PLK-1磷酸化功能，与Scripps研究研究所生理蛋白质组学中心合作，我们通过使用体外激酶分析和分析质谱法绘制了KNL-1上的PLK-1目标位点。在我们提交分析的3个样本中，多次鉴定了6个磷岩，这表明它们是PLK-1磷酸化的KNL-1主要部位。现在，我们正在研究这些磷酸化在控制甲状腺丝网胚胎中的有丝分裂进展和染色体分离中的作用。