Ultrasound generates both compressive and shear mechanical forces in soft tissues. However, the specific mechanisms by which these forces activate cellular processes remain unclear. Here we show that low-intensity focused ultrasound can activate the mechanosensitive RET signalling pathway. Specifically, in mouse colon tissues ex vivo and in vivo, focused ultrasound induced RET phosphorylation in colonic crypts cells, which correlated with markers of proliferation and stemness when using hours-long insonication. The activation of the RET pathway is non-thermal, is linearly related to acoustic pressure and is independent of radiation-force-induced shear strain in tissue. Our findings suggest that ultrasound could be used to regulate cell proliferation, particularly in the context of regenerative medicine, and highlight the importance of adhering to current ultrasound-safety regulations for medical imaging.