SIGNAL TRANSDUCTION OF CELL DENSITY SENSING IN DICTYOSTELIUM DISCOIDEUM

盘基网柄菌细胞密度传感的信号转导

• 批准号: 6313787
• 负责人: DERRICK T BRAZILL
• 金额: $ 5.95万
• 依托单位: HUNTER COLLEGE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6313787
• 关键词: Dictyostelium; G protein; biological signal transduction; biosensor device; cell differentiation; cell growth regulation; cell migration; cell morphology; cell proliferation; cyclic AMP; enzyme activity; guanosinetriphosphatases; immunoprecipitation; phosphorylation; protein kinase C; protein protein interaction; protein purification; protein structure function; receptor binding; transcription factor; western blottings

The ability of mammalian cell to sense the density of the cells around it plays an important role in cellular growth and differentiation. Without such an ability, a developing embryo would be unable to properly proportion its cells into different tissue types. Defects in cell density sensing may also be involved in cancer. Unfortunately, studying cell density sensing in mammalian cells is difficult due to their genetic intractability. However, the simple eukaryote Dictyostelium discoideum is an excellent genetic organism for the study of such a phenomenon. Dictyostelium cells monitor the density of starving cells by sensing the levels of CMF, a protein secreted by starving cells. When there is a sufficiently high density of starving cells, as indicated by a high extracellular concentration of CMF, the cells use relayed pulses of cAMP as a chemoattractant to allow them to aggregate and undergo development. cAMP binds to its receptor, cAR1, and activates a G protein signaling cascade culminating in the regulation of developmentally important genes. However, in the absence of CMF , while cAMP still binds to and activates its receptor, it does not activate downstream signaling components. CMF accomplishes the regulation by controlling the activity of the G protein associated with cAR1. This proposal describes experiments that will characterize the ability of CMF to regulate a G protein and delineate the proteins involved in this regulation. Specifically, the ability of CMF to affect the phosphorylation status of cAR1, the interaction cAR1 and its G protein, and the localization of known signaling proteins to the membrane will be examined. In addition, molecules that are involved in the regulation of the GTPase activity of the G protein will be identified.

哺乳动物细胞感知周围细胞密度的能力在细胞生长和分化中起着重要作用。没有这种能力，发育的胚胎将无法将其细胞正确比例为不同的组织类型。细胞密度传感的缺陷也可能参与癌症。不幸的是，由于哺乳动物细胞的细胞密度感测，由于其遗传性疾病性很难。然而，简单的真核生物dictyostelium discoideum是研究这种现象的出色遗传生物。 Dictyostelium细胞通过感测CMF的水平来监测饥饿细胞的密度，CMF的水平是由饥饿细胞分泌的蛋白质。当有足够高的饥饿细胞密度（如高细胞外浓度CMF所示）时，细胞使用cAMP的继电器脉冲作为化学吸引剂，使它们可以聚集并进行发育。 CAMP与其受体CAR1结合，并激活G蛋白信号传导级联反应在调节发育重要基因的调节中。但是，在没有CMF的情况下，cAMP仍然与其受体结合并激活其受体，但它不会激活下游信号传导成分。 CMF通过控制与CAR1相关的G蛋白的活性来实现调节。该建议描述了将表征CMF调节G蛋白并描述涉及该调节的蛋白质的能力的实验。具体而言，将检查CMF影响CAR1的磷酸化状态，相互作用CAR1及其G蛋白的能力，以及已知信号蛋白在膜上的定位。另外，将鉴定参与GTPase活性的分子。