Context-dependent inhibitory roles of RhoA in 3D invasive cell migration within the extracellular matrix

Cell migration is fundamental to both physiological and pathological processes, including cancer progression. This study investigates the role of the small GTPase RHOA in invasive cell migration within diverse 3D extracellular matrix (ECM) environments using non-cancerous HEK293, pancreatic cancer PANC-1, and breast cancer MDA-MB-231 cells. Spheroid invasion assays showed that RHOA loss enhanced migration in HEK293 and PANC-1 cells cultured in Geltrex but not in type I collagen. In contrast, RHOA deletion had little effect on MDA-MB-231 migration in either ECM. Enhanced migration in RHOA-deficient HEK293 cells required protein phosphatase PTP1B and the small GTPases RAC and CDC42. Unexpectedly, while RHOA knockout increased 3D migration, it reduced pancreatic tumor progression in mice. These findings reveal that RHOA regulates cell invasion in a manner dependent on ECM composition and cellular context, highlighting its complex, context-specific roles and potential as a therapeutic target in cancer.