Environmentally dependent and independent control of cell shape determination by Rho GTPase regulators in melanoma

In order to invade 3D tissues, cancer cells dynamically change cell morphology in response to geometric and mechanical cues in the environment. But how cells determine their shape in 3D versus 2D environments is poorly understood. Studying 2D versus 3D single cell shape determination has historically been technically difficult due to the lack of methodologies to directly compare the two environments. We developed an approach to study cell shape in 2D versus 3D by measuring cell shape at different depths in collagen using stage-scanning oblique plane microscopy (ssOPM). We find characteristic shape changes occur in melanoma cells depending on whether a cell is attached to a 2D surface or 3D environment, and that these changes can be modulated by Rho GTPase regulatory proteins. Our data suggest that regulation of cell protrusivity undergoes a ‘switch’ of control between different Rho GTPase regulators depending on the physical microenvironment. ### Competing Interest Statement Chris Dunsby has a licensed granted patent on OPM. * LSFM : light sheet fluorescence microscopy OPM : oblique plane microscopy PSF : point spread function ssOPM : stage scanning oblique plane microscopy