The Extracellular Matrix Regulates Invasion in Fusion-Negative Rhabdomyosarcoma via YAP-PIEZO1 Signaling Axis

Background Fusion-negative rhabdomyosarcoma (FNRMS) represents the most prevalent subtype of rhabdomyosarcoma, the most common pediatric soft tissue sarcoma. Although its invasion is a leading cause of recurrence and poor prognosis, its underlying mechanism remains unclear. We investigated how extracellular matrix (ECM) density regulates FNRMS progression via mechano-transduction. Methods We used three-dimensional spheroid invasion assays with RD and SMS-CTR cells embedded in varying collagen concentrations. Mechanistic insights were gained through immunofluorescence, ChIP-seq re-analysis, Fluo-4 calcium live cell imaging, and pharmacological inhibition of the YAP–PIEZO1 axis. Results High ECM density significantly enhanced invasive spreading, correlating with increased YAP nuclear localization. YAP overexpression was sufficient to promote invasion, while its inhibition attenuated the matrix-enhanced phenotype. We identified PIEZO1 as a direct transcriptional target of the YAP. High ECM density stimulated PIEZO1-dependent calcium influx, which was required for invasion. Furthermore, elevated PIEZO1 expression was significantly associated with poorer overall survival in FNRMS patients. Targeting YAP or ROCK signaling effectively suppressed both calcium flux and invasion. Conclusions Our findings establish a YAP–PIEZO1 axis linking ECM density to FNRMS invasion. This mechanosensitive pathway represents a potential therapeutic vulnerability in aggressive FNRMS. Simple Summary Fusion-negative rhabdomyosarcoma (FNRMS) is the prevalent subtype of rhabdomyosarcoma, the most common pediatric soft tissue sarcoma. Its invasion often leads to recurrence and poor prognosis. This study investigates how the density of the extracellular matrix (ECM), surrounding cancer cells in the tissue, influences FNRMS invasion. We found that high ECM density activates the protein YAP, which directly triggers the expression of the mechanical sensor PIEZO1. This sensor allows calcium to enter the cells, providing a signal that facilitates invasion. Pharmacological inhibition of this axis successfully reduced the invasive potential, highlighting a novel therapeutic vulnerability for FNRMS patients.