Neuronal paxillin and drebrin mediate BDNF-induced force transduction and growth cone turning in a soft tissue-like environment

Soft tissue environments govern neuronal morphogenesis. However, the precise molecular mechanisms underlying chemotropism-directed axonal growth cone movement in extremely soft environments remain unclear. Here, we show that drebrin, a growth cone T-zone protein, modulates growth cone turning in response to BDNF coated on a soft substrate. Structurally, axonal growth cones of rodent hippocampal neurons grown on 0.1 kPa hydrogels possess an expanded T-zone in which drebrin is highly integrated with both F-actin and microtubules. Biochemically, we identify paxillin as interacting with drebrin in cells grown on 0.1 kPa hydrogels but not on glass coverslips. When grown on 0.1 kPa substrates, growth cones asymmetrically exposed to BDNF-bound stripes exhibit enhanced paxillin/drebrin interaction on the side facing the stripes, an activity that is PKA- and AAK1-dependent, but independent of Src kinase. Functionally, we show that BDNF-induced growth cone turning and force generation on soft substrates require drebrin phosphorylation and paxillin/drebrin association.

软组织微环境调控神经元形态发生。然而，极端柔软环境中由趋化性介导的轴突生长锥运动背后的确切分子机制仍不明晰。本研究发现，生长锥T区蛋白脑发育调节蛋白（drebrin）可调控软基质包被的脑源性神经营养因子（BDNF）诱导的生长锥转向。结构层面，在0.1 kPa水凝胶上培养的啮齿类海马神经元的轴突生长锥具有扩张的T区，其中drebrin可与纤维状肌动蛋白（F-actin）及微管紧密结合。生化层面，我们证实，在0.1 kPa水凝胶上培养的细胞中，桩蛋白（paxillin）可与drebrin发生相互作用，而在玻璃盖玻片上培养的细胞中则无此相互作用。当在0.1 kPa基质上培养时，不对称暴露于BDNF包被条纹的生长锥，在朝向条纹的一侧会增强桩蛋白与drebrin的相互作用，该过程依赖于蛋白激酶A（PKA）与AP2关联激酶1（AAK1），但不依赖于Src激酶（Src kinase）。功能层面，我们证实，软基质上BDNF诱导的生长锥转向与力产生过程，需要drebrin的磷酸化以及桩蛋白与drebrin的相互结合。