Importance of non-selective cation channel TRPV4 interaction with cytoskeleton and their reciprocal regulations in cultured cells.
Importance of non-selective cation channel TRPV4 interaction with cytoskeleton and their reciprocal regulations in cultured cells.
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DOI：
10.1371/journal.pone.0011654
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发表时间：
2010-07-19
期刊：
影响因子：
3.7
通讯作者：
Hucho T
中科院分区：
文献类型：
--
作者：
Goswami C;Kuhn J;Heppenstall PA;Hucho T
TRPV 4和细胞骨架都被报道影响细胞机械敏感过程以及机械痛觉过敏的发展。TRPV 4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DRG神经元和DRG神经元衍生的F-11细胞的形态学变化，以及钙离子成像，从生化、细胞生物学和功能上研究了TRPV 4与细胞骨架成分的相互作用。我们发现TRPV 4与微管蛋白、肌动蛋白和神经丝蛋白以及伤害感受分子PKCε和CamKII发生物理相互作用。TRPV 4的C-末端足以与微管蛋白和肌动蛋白直接相互作用，无论是其可溶形式还是其聚合形式。肌动蛋白和微管蛋白竞争结合。与TRPV 4的相互作用稳定微管，即使在体外解聚条件下。因此，在细胞系统中，TRPV 4与肌动蛋白和微管富集结构在膜下区域共定位。TRPV 4的表达和活化均诱导显著的形态学变化，影响非神经元细胞、DRG神经元衍生的F11细胞以及IB 4阳性DRG神经元中的板状伪足、丝状伪足、生长锥和神经突结构。TRPV 4和细胞骨架的功能相互作用是相互的，因为微管稳定剂Taxol通过TRPV 4减少Ca 2+内流。TRPV 4作为微管和肌动蛋白的调节剂。反过来，我们描述了微管动力学是TRPV 4活性的重要调节剂。TRPV 4形成包含细胞骨架蛋白和调节激酶的超分子复合物。因此，它可以整合各种细胞内第二信使和信号级联的信号传导，以及细胞骨架动力学。本研究指出，非选择性阳离子通道与细胞骨架之间存在多层次的交互作用。这些交互作用可能有助于我们理解在癌症患者中常见的紫杉醇诱导的神经病理性疼痛发展的分子基础。
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 PKCε 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.
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