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，超软环境中趋化性引导的轴突生长锥运动背后的确切分子机制仍未阐明。本研究发现，生长锥T区蛋白drebrin（drebrin）可调控软底物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0.1kPa水凝胶上培养的啮齿类海马神经元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）和微管高度整合。生化层面，我们发现在0.1kPa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0.1kPa底物上培养时，不对称暴露于包被有BDNF的条纹区域的生长锥，其朝向条纹的一侧会出现桩蛋白与drebrin相互作用增强的现象，该过程依赖于蛋白激酶A（PKA）和AAK1（AAK1），但不依赖于Src激酶（Src kinase）。功能层面，我们证实软底物上BDNF诱导的生长锥转向与力产生过程，均依赖于drebrin的磷酸化以及桩蛋白与drebrin的结合。