PATHWAYS OF RECEPTOR SIGNALING AND CHEMOTAXIS

受体信号传导和趋化性的途径

• 批准号: 7593426
• 负责人: ALAN R KIMMEL
• 金额: $ 46.55万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7593426
• 关键词: Accounting; Actins; Adenylate Cyclase; Binding; Biological Process; CAR receptor; Cell Polarity; Cell Surface Receptors; Cells; Chemicals; Chemotactic Factors; Chemotaxis; Complex; Condition; Cyclic AMP; Cyclic AMP Receptors; Cyclic AMP-Dependent Protein Kinases; Cytoskeleton; Development; Dictyostelium; Eukaryota; Eukaryotic Cell; Family; Feedback; GTP-Binding Proteins; Gene Expression; Growth and Development function; Heterotrimeric GTP-Binding Proteins; Individual; Ligand Binding; Ligands; MAPK1 gene; Mammalian Cell; Mediating; Mitogen-Activated Protein Kinases; Neurotransmitters; Pathway interactions; Phosphorylation; Phosphotransferases; Physiological Processes; Production; Range; Receptor Signaling; Regulation; Serine; Signal Pathway; Signal Transduction; System; Threonine; Transmembrane Domain; Variant; Work; chemokine; chemokine receptor; extracellular; morphogens; neutrophil; polarized cell; receptor; response

Chemotactic cells orient and move directionally in very shallow chemical gradients. As cells polarize, distinct structural and signaling components become spatially restricted to the leading edge or rear of cells. Feedback loops downstream of receptor signaling integrate both activating and inhibiting pathways to establish cell polarity within such gradients. While much effort has focused on defining activating pathways, inhibitory networks have been largely unexplored. Many receptor-mediated pathways in eukaryotes adapt/deactivate to persistent stimulation. While the MAP kinase ERK2 in Dictyostelium has been assumed to adapt to continuous engagement of cAMP with its 7-transmembrane, cell surface receptor CAR, we have shown, to the contrary, that ERK2 remains active under such conditions. The upstream phosphorylation pathway that is responsible for ERK2 activation transiently responds to CAR stimulation, whereas ERK2 de-phosphorylation (deactivation) is inhibited by continuous stimulation. We argue that the net result is persistently active ERK2 when the extracellular cAMP concentration is constant and that the oscillating production/destruction of secreted cAMP in chemotaxing cells accounts for the observed oscillatory activity of ERK2. We also showed that CAR-dependent pathways that control ERK2 activation/deactivation function independently of G proteins and of ligand-induced production of intracellular cAMP and the consequent activation of PKA. This regulation enables ERK2 to function both in an oscillatory manner, critical for chemotaxis, and in a persistent manner, necessary for gene expression, as secreted cAMP increases during later development. This work redefines mechanisms of ERK2 regulation by CAR signaling in Dictyostelium and establishes new implications for control of signal-relay during chemotaxis. 7-TMRs activate multiple downstream signaling cascades via heterotrimeric G protein-dependent and -independent pathways and control a wide range of biological processes. Upon ligand binding, 7-TMRs become phosphorylated at cytoplasmic serine and threonine residues by specific receptor kinases, functioning to uncouple heterotrimeric G protein pathways from receptor signaling and to activate G protein-independent pathways. In Dictyostelium, CAR signaling regulates chemotaxis and both G protein-dependent and -independent signaling cascades. Since the CARs become phosphorylated upon cAMP binding, we analyzed the ability of cells that only express non-phosphorylatable CAR variants to respond to cAMP. These cells were defective in adenylyl cyclase adaptation and in cell polarization and chemotaxis. While receptor phosphorylation is known to uncouple certain G protein-mediated signaling pathways from 7-TMRs in mammalian cells, our studies are the first to show that such mechanisms also function in pathways that regulate chemotaxis.

趋化细胞在非常浅的化学梯度中定向和移动。当细胞极化时，不同的结构和信号成分在空间上被限制在细胞的前缘或后部。受体信号传导下游的反馈环路整合了激活和抑制途径，以在这种梯度内建立细胞极性。虽然很多努力都集中在定义激活途径上，但抑制网络基本上尚未被探索。 真核生物中的许多受体介导的途径会适应/失活持续刺激。虽然盘基网柄菌中的 MAP 激酶 ERK2 被认为能够适应 cAMP 与其 7 次跨膜细胞表面受体 CAR 的持续结合，但我们已经证明，相反，ERK2 在这种条件下仍然保持活性。负责 ERK2 激活的上游磷酸化途径会短暂响应 CAR 刺激，而 ERK2 去磷酸化（失活）会受到持续刺激的抑制。我们认为，当细胞外 cAMP 浓度恒定时，最终结果是 ERK2 持续活跃，并且趋化细胞中分泌的 cAMP 的振荡产生/破坏解释了观察到的 ERK2 振荡活性。我们还表明，控制 ERK2 激活/失活的 CAR 依赖性途径独立于 G 蛋白和配体诱导的细胞内 cAMP 的产生以及随后的 PKA 激活。这种调节使 ERK2 能够以对趋化性至关重要的振荡方式发挥作用，并以基因表达所必需的持久方式发挥作用，因为在后期发育过程中分泌的 cAMP 会增加。这项工作重新定义了盘基网柄菌中 CAR 信号传导对 ERK2 的调节机制，并为趋化过程中信号传递的控制建立了新的含义。 7-TMR 通过异源三聚体 G 蛋白依赖性和非依赖性途径激活多个下游信号级联，并控制广泛的生物过程。配体结合后，7-TMR 通过特定受体激酶在细胞质丝氨酸和苏氨酸残基处磷酸化，从而将异源三聚体 G 蛋白途径与受体信号传导分离并激活不依赖于 G 蛋白的途径。在盘基网柄菌中，CAR 信号传导调节趋化性以及 G 蛋白依赖性和独立性信号级联。由于 CAR 在 cAMP 结合后被磷酸化，因此我们分析了仅表达不可磷酸化 CAR 变体的细胞响应 cAMP 的能力。这些细胞在腺苷酸环化酶适应、细胞极化和趋化性方面存在缺陷。虽然已知受体磷酸化可将某些 G 蛋白介导的信号通路与哺乳动物细胞中的 7-TMR 解偶联，但我们的研究首次表明此类机制也在调节趋化性的通路中发挥作用。