Signal Transduction in Pain and Mental Retardation, Department for Molecular Human Genetics Max Planck Institute for Molecular Genetics, Berlin, Germany.
PLoS One. 2010 Jul 19;5(7):e11654. doi: 10.1371/journal.pone.0011654.
TRPV4 and the cellular cytoskeleton have each been reported to influence cellular mechanosensitive processes as well as the development of mechanical hyperalgesia. If and how TRPV4 interacts with the microtubule and actin cytoskeleton at a molecular and functional level is not known.
We investigated the interaction of TRPV4 with cytoskeletal components biochemically, cell biologically by observing morphological changes of DRG-neurons and DRG-neuron-derived F-11 cells, as well as functionally with calcium imaging. We find that TRPV4 physically interacts with tubulin, actin and neurofilament proteins as well as the nociceptive molecules PKCepsilon and CamKII. The C-terminus of TRPV4 is sufficient for the direct interaction with tubulin and actin, both with their soluble and their polymeric forms. Actin and tubulin compete for binding. The interaction with TRPV4 stabilizes microtubules even under depolymerizing conditions in vitro. Accordingly, in cellular systems TRPV4 colocalizes with actin and microtubules enriched structures at submembranous regions. Both expression and activation of TRPV4 induces striking morphological changes affecting lamellipodial, filopodial, growth cone, and neurite structures in non-neuronal cells, in DRG-neuron derived F11 cells, and also in IB4-positive DRG neurons. The functional interaction of TRPV4 and the cytoskeleton is mutual as Taxol, a microtubule stabilizer, reduces the Ca2+-influx via TRPV4.
TRPV4 acts as a regulator for both, the microtubule and the actin. In turn, we describe that microtubule dynamics are an important regulator of TRPV4 activity. TRPV4 forms a supra-molecular complex containing cytoskeletal proteins and regulatory kinases. Thereby it can integrate signaling of various intracellular second messengers and signaling cascades, as well as cytoskeletal dynamics. This study points out the existence of cross-talks between non-selective cation channels and cytoskeleton at multiple levels. These cross talks may help us to understand the molecular basis of the Taxol-induced neuropathic pain development commonly observed in cancer patients.
瞬时受体电位香草酸亚型 4（TRPV4）和细胞骨架都被报道影响细胞机械敏感过程以及机械性痛觉过敏的发展。TRPV4 是否以及如何在分子和功能水平上与微管和肌动蛋白细胞骨架相互作用尚不清楚。
我们通过观察背根神经节神经元和背根神经节神经元衍生的 F-11 细胞的形态变化，以及通过钙成像进行功能研究，从生化和细胞生物学方面研究 TRPV4 与细胞骨架成分的相互作用。我们发现 TRPV4 与微管蛋白、肌动蛋白和神经丝蛋白以及痛觉分子 PKCepsilon 和 CamKII 发生物理相互作用。TRPV4 的 C 端足以与微管蛋白和肌动蛋白直接相互作用，无论是可溶性形式还是聚合形式。肌动蛋白和微管蛋白竞争结合。在体外解聚条件下，这种相互作用稳定微管。因此，在细胞系统中，TRPV4 与富含亚膜区的肌动蛋白和微管富集结构共定位。TRPV4 的表达和激活都会引起非神经元细胞、背根神经节衍生的 F11 细胞以及 IB4 阳性背根神经节神经元中明显的形态变化，影响片状伪足、丝状伪足、生长锥和神经突结构。TRPV4 和细胞骨架的功能相互作用是相互的，因为紫杉醇是一种微管稳定剂，可减少 TRPV4 引起的 Ca2+内流。
TRPV4 既是微管的调节剂，也是肌动蛋白的调节剂。反过来，我们描述了微管动力学是 TRPV4 活性的重要调节剂。TRPV4 形成一个包含细胞骨架蛋白和调节激酶的超分子复合物。由此，它可以整合各种细胞内第二信使和信号级联的信号以及细胞骨架动力学。这项研究指出了非选择性阳离子通道和细胞骨架在多个层面上存在串扰。这些串扰可能有助于我们理解紫杉醇引起的癌症患者中常见的神经性疼痛发展的分子基础。
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