Distinct Mechanosensing of Human Neural Stem Cells on Extremely Limited Anisotropic Cellular Contact

Human neural stem cells (hNSCs) can alter their fate choice in response to the biophysical cues provided during development. In particular, it has been reported that the differentiation of neural stem cells (NSCs) is enhanced by anisotropic contact, which facilitates focal adhesion (FA) formation and cytoskeletal organization. However, a biomolecular mechanism governing how the cells process the biophysical cues from these anisotropic geometries to their fate commitment is still poorly understood due to the limited availability of geometrical diversities (contact width above 50 nm) applicable to cell studies. Here, we firstly demonstrate that the biomolecular mechanism for enhanced neurogenesis on an anisotropic nanostructure is critically dependent on the resolution of a contact feature. We observed a totally different cellular response to anisotropic geometries by first utilizing a high-resolution nanogroove (HRN) structure with an extremely narrow contact width (15 nm). The width scale is sufficiently low to suppress the integrin clustering and enable us to elucidate how the contact area influences the neurogenesis of hNSCs at an aligned state. Both the HRN and control nanogroove (CN) pattern with a contact width of 1 μm induced the spontaneous topographic alignment of hNSCs. However, intriguingly, the focal adhesion (FA) formation and cytoskeletal reorganization were substantially limited on the HRN, although the cells on the CN showed enhanced FA formation compared with flat surfaces. In particular, the hNSCs on the HRN surface exhibited a strikingly lower fraction of nuclear yes-associated protein (YAP) than on the CN surface, which was turned out to be regulated by Rho GTPase in the same way as the cells sense the mechanical properties of the environment. Considering the previously reported role of YAP on neurogenesis, our finding newly substantiates that YAP and Rho GTPase also can be transducers of hNSCs to process topographical alternation to fate decision. Furthermore, this study with the unprecedented high-resolution nanostructure suggests a novel geometry sensing model where the functional crosstalk between YAP signaling and Rho GTPase integrally regulate the fate commitment of the hNSCs.