Molecular Mechanisms of Signal Transduction Involving Light, Redox and Transmembrane Complexes

Crane研究小组研究信号转导机制，总体目标是理解行为 分子水平。这种理解将通过定义KEY的结构和动态来实现 在两个系统中协调基因表达和跨膜信号的大分子复合体 依靠高度合作的相互作用来响应光、氧化还原和化学环境。第一，细菌 趋化性，关注原核细胞对外界刺激物的运动。趋化性是一种 了解跨膜通讯、细胞内信息传递和运动性。重要的是 导致霍乱、胃癌和莱姆病等疾病的许多人类病原体依赖于趋化作用 确定感染。化学趋化性的基础感觉器官，以下称为“化学小体”，显示 惊人的灵敏度、动态范围和基本的分子记忆。在化学小体中，受体， 组氨酸激酶(CheA)和偶联蛋白组装成一个特定的结构，其细节只是 正在浮现。这一建议继续努力了解化学小体组装，化学受体构象 通过重组受体阵列发出信号，并最终调控CHEA。化学小体产量 调节大自然完美的纳米机器--鞭毛马达。开关复合体的超微结构 将定义电机内的扭矩产生、方向切换和响应 化学小体信号。第二个系统，真核生物生物钟，包括细胞自主计时。 使新陈代谢达到昼夜周期的装置。时钟由转录-翻译组成 抑制蛋白抑制自身基因转录激活子的反馈环(TTFL)。 光通过刺激直接照射在TTFL上的光传感器来引入时钟相位。在人类身上， 时钟功能异常会导致精神疾病(睡眠障碍、抑郁、躁狂)、细胞生长失调 (癌症)和代谢缺陷(糖尿病和肥胖症)。这个项目提出了结构性和机械性 对高等生物体中常见的关键抑制物和光设置活动的研究。 将对真菌(粗枝脉孢菌)和苍蝇的昼夜节律蛋白进行生物物理研究 (黑腹果蝇)。这两种模式生物都提供了遗传系统和行为分析来进行探索 机械论洞察力的生物学相关性。一套免费的技术，包括X光 结晶学、小角X射线散射、光谱学、低温电子显微镜和脉冲偶极 将应用电子自旋共振波谱(PDS)来实现这些目标。对于PDS，整合的新战略 将开发和部署基于氮氧化物、黄素、核苷酸和金属离子的自旋探针。总的来说， 该计划旨在提供对细菌趋化性的感知和反应的分子理解，以及 通过生物物理方法的协同应用研究真核生物的昼夜节律。

The Crane group studies mechanisms of signal transduction with the overall goal of understanding behavior at the molecular level. This understanding will be achieved by defining the structure and dynamics of key macromolecular complexes that coordinate gene expression and transmembrane signaling in two systems that rely on highly cooperative interactions to respond to light, redox and chemical environment. The first, bacterial chemotaxis, concerns the motion of prokaryotic cells toward external stimulants. Chemotaxis is a paradigm for understanding transmembrane communication, intracellular information transfer, and motility. Importantly, many human pathogens that cause diseases such as cholera, gastric cancer and lyme rely on chemotaxis to establish infection. The sensory apparatus underlying chemotaxis, hereafter called “the chemosome”, displays amazing sensitivity, dynamic range and a rudimentary molecular memory. In the chemosome, receptors, histidine kinases (CheA) and coupling proteins assemble into a specific architecture, whose details are just emerging. This proposal continues efforts to understand chemosome assembly, chemoreceptor conformational signaling, and ultimately, CheA regulation through restructuring of the receptor arrays. Chemosome output modulates Nature's consummate nanomachine – the flagella motor. The ultrastructure of the switch complex within the motor will be defined to understand torque generation, direction switching and response to chemosome signals. The second system, eukaryotic circadian clocks, comprises cell-autonomous timing devices that pace metabolism to the diurnal cycle. Clocks are composed of transcriptional-translational feedback loops (TTFLs) within which repressor proteins inhibit the transcriptional activators of their own genes. Light entrains the clock phase by stimulating photosensors that impinge directly on the TTFLs. In humans, aberrant clock function causes mental illness (sleep disorders, depression, mania), cell growth deregulation (cancer) and metabolic defects (diabetes and obesity). This project proposes structural and mechanistic investigations of the key repressor and light-setting activities common to clocks in higher organisms. Biophysical studies will be conducted on the circadian proteins of fungi (Neurospora crassa) and flies (Drosophila melanogaster). Both model organisms provide genetic systems and behavioral assays to probe the biological relevance of mechanistic insights. A complimentary set of techniques including X-ray crystallography, small-angle X-ray scattering, optical spectroscopy, cryo-electron microscopy and pulse-dipolar ESR spectroscopy (PDS) will be applied to accomplish these goals. For PDS, new strategies for incorporating spin probes based on nitroxides, flavins, nucleotides, and metal ions will be developed and deployed. Overall, this program aims to provide a molecular understanding for sensing and response in bacterial chemotaxis and eukaryotic circadian rhythms through the synergistic application of biophysical methods.