P1-SPATIAL REGULATION OF PROTEIN KINASE A SIGNALING DURING GROWTH CONE GUIDANCE

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。 在发育过程中，细胞外的指导线索调节肌动蛋白细胞骨架动力学在神经生长锥，以控制轴突的生长和轨迹向正确的突触目标。在这些线索中有netrins，一个家族的分泌分子能够化学吸引，化学排斥，并调节分支行为。对netrin信号传导的应答主要通过DCC（结直肠癌中的神经递质）受体家族介导。描绘DCC下游的信号和它们与细胞骨架动力学耦合的机制对于我们理解netrin介导的发育事件至关重要。我们以前已经表明，cAMP依赖性蛋白激酶（PKA）在趋化性细胞运动过程中的空间调节和PKA的亚细胞锚定是必需的有效的趋化性。我们的初步数据表明DCC的表达以配体依赖的方式促进PKA的活化。这种活性与Mena/VASP家族的细胞骨架调节剂的磷酸化相关，这种作用通过破坏PKA锚定而消除，表明涉及特定的含有PKA的信号传导复合物。我们还观察到PKA和DCC的免疫共沉淀被PKA锚定的抑制所破坏。这些数据支持一个假设，其中协会，并激活，PKA DCC参与耦合netrin/DCC信号转导细胞骨架调节轴突导向。我们将通过（1）阐明DCC和PKA之间相互作用的性质，（2）研究DCC与PKA相关和通过PKA信号传导的分子后果，以及（3）研究DCC/PKA信号传导对神经元形态，动力学和指导的功能后果来测试这一点。

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. During development, extracellular guidance cues regulate actin cytoskeletal dynamics in neuronal growth cones to control axon outgrowth and trajectory towards proper synaptic targets. Among these cues are the netrins, a family of secreted molecules capable of chemoattraction, chemorepulsion, and regulation of branching behavior. The response to netrin signaling is mediated principally through the DCC (Deleted in Colorectal Cancers) receptor family. Delineation of the signals downstream of DCC and the mechanisms through which they couple to cytoskeletal dynamics is fundamentally important to our understanding of netrin-mediated developmental events. We have previously shown that the cAMP-dependent protein kinase (PKA) is spatially regulated during chemotactic cell movement and that subcellular anchoring of PKA is required for efficient chemotaxis. Our preliminary data show that expression of DCC promotes activation of PKA in a ligand-dependent fashion. This activity correlates with phosphorylation of the Mena/VASP family of cytoskeletal regulators, an effect abrogated by disruption of PKA anchoring, suggesting the involvement of a specific PKA-containing signaling complex. We have also observed that co-immunoprecipitation of PKA and DCC is disrupted by inhibition of PKA anchoring. These data support a hypothesis in which association with, and activation of, PKA by DCC is involved in coupling netrin/DCC signaling to cytoskeletal regulation during axon guidance. We will test this by (1) elucidating the nature of the interaction between DCC and PKA, (2) investigating the molecular consequences of DCC association with and signaling through PKA, and (3) investigating the functional consequences of DCC/PKA signaling on neuronal morphology, dynamics, and guidance.