β1-Integrin cytoskeletal signaling regulates sensory neuron response to matrix dimensionality.

Neuronal differentiation, pathfinding and morphology are directed by biochemical cues that in vivo are presented in a complex scaffold of extracellular matrix. This microenvironment is three-dimensional (3D) and heterogeneous. Therefore, it is not surprising that more physiologically-relevant cellular responses are found in 3D culture environments rather than on two-dimensional (2D) flat substrates. One key difference between 2D and 3D environments is the spatial arrangement of cell-matrix interactions. Integrins and other receptor proteins link the various molecules presented in the extracellular environment to intracellular signaling cascades and thus influence a number of neuronal responses including the availability and activation of integrins themselves. We have previously reported that a 3D substrate induces an important morphological transformation of embryonic mouse dorsal root ganglion (DRG) neurons. Here, we investigate the hypothesis that β1-integrin signaling via focal adhesion kinase (FAK) and the RhoGTPases Rac and Rho influences neuronal morphology in 2D vs 3D environments. We report that β1-integrin activity and FAK phosphorylation at tyrosine 397 (FAKpY397) are linked to neuronal polarization as well as neurite outgrowth and branching. Rac and Rho expression are decreased in 3D vs 2D culture but not correlated with β1-integrin function. These results suggest that proper β1-integrin activity is required for the elaboration of physiologic DRG morphology and that 3D culture provides a more appropriate milieu to the mimic in vivo scenario. We propose that neuronal morphology may be directed during development and regeneration by factors that influence how β1-integrin, FAK and RhoGTPase molecules integrate substrate signals in the 3D microenvironment.