2 These authors made an equal contribution. Basement membrane molecules such as laminin are important structural components of the skin 1-4 , but also serve as substrates for sensory neurons of the dorsal root ganglia (DRG) to grow in culture 5 . The main function of sensory neurons innervating the skin is to detect and relay relevant sensory stimuli, in particular mechanical stimuli 6 . It has long been known that sensory neurons with a nociceptive function (detecting potentially harmful stimuli) can have their endings in the epidermis 7-9 whereas mechanoreceptor endings (touch receptors) reside exclusively in the dermal layer [9][10] . Interestingly, the matrix environments of the epidermis and the dermis are very distinctive 11 . We showed that mechanosensitive currents required for touch receptor function depend on the presence of a protein tether which may function to couple mechanosensitive channels to a laminin-containing matrix 12 . The tether protein is not required for the mechanosensitivity of most nociceptive sensory neurons. Here we set out to address the idea that sensory mechanotransduction might be modulated by distinct matrix components made by different types of skin cells in different skin layers. We show that epidermal keratinocytes produce a matrix that is non-permissive for mechanotransduction and identify the factor responsible as laminin-332 (formerly known as laminin-5). Laminin matrices doped with small amounts of laminin-332 have a dramatically altered network structure that is non-permissive for tether attachment.We demonstrate a spatially restricted loss of mechanotransduction in neurite segments connected to laminin-332-containing matrices. Mutations in all three genes coding the trimeric laminin-332 protein complex can cause epidermolysis bullosa, a severe inherited skin blistering disease 1,3 . Human keratinocytes that produce a laminin-332 free matrix have no inhibitory activity on mechanotransduction. We have also discovered an activity of laminin-332 matrix in inhibiting sensory axon bifurcation. Our results reveal novel mechanisms whereby permissive and non-permissive substrates can spatially coordinate mechanotransduction in distinct domains within a single neuron. Results Keratinocyte matrix is suppresses mechanotransductionUsing whole-cell, patch-clamp techniques we directly recorded mechanosensitive currents in cultured sensory neurons [12][13][14][15][16][17][18][19][20] . We first asked whether co-culture of sensory neurons with different cellular components of the skin can modulate the activity of mechanosensitive currents. When sensory neurons are cultured on a laminin substrate, standardly-derived from Engelbreth-Holm-Swarm cells (EHS matrix, henceforth referred to as laminin), more than 90% of the cells exhibit a mechanosensitive current evoked using a small (~740 nm displacement) stimulus to the neurite 12, 14-15 . At least three types of mechanosensitive current can be measured in sensory neurons, classified according to their inactivation time constant τ 1 , rap...