An object of the present invention is to provide a Group III nitride semiconductor multilayer structure having a smooth surface and exhibiting excellent crystallinity, which multilayer structure employs a low-cost substrate that can be easily processed. Another object is to provide a Group III nitride semiconductor light-emitting device comprising the multilayer structure.The inventive Group III nitride semiconductor multilayer structure comprises a substrate; an AlxGa1-xN (0≦x≦1) buffer layer which is provided on the substrate and has a columnar or island-like crystal structure; and an AlxInyGa1-x-yN (0≦x≦1, 0≦y≦1, 0≦x+y≦1) single-crystal layer provided on the buffer layer, wherein the substrate has, on its surface, non-periodically distributed grooves having an average depth of 0.01 to 5 μm.