PDGFRα+/Integrin α2+ Fibroblasts Orchestrate Tumor Budding in Oral Squamous Cell Carcinoma via Mechano-Metabolic Symbiosis: E-Cadherin/Integrin α2β1 Adhesion and Mitochondrial Transfer.

Tumor budding (TB), defined as single tumor cells or small clusters (≤4 cells) at the invasive front, is a strong adverse prognostic feature in oral squamous cell carcinoma (OSCC). However, the stromal regulators and molecular mechanisms that drive TB remain unclear. Here, we developed a tumor-budding organoid (TBO) model that faithfully recapitulates the OSCC tumor-stroma interface and budding dynamics. By integrating this model with single-cell RNA sequencing (scRNA-seq), in situ RNA sequencing (isRNA-seq), and functional perturbations, we identify platelet-derived growth factor receptor alpha+/integrin α2+ (PDGFRα+/integrin α2+) cancer-associated fibroblasts (CAFs) as the key stromal subset promoting OSCC budding. OSCC cells recruit PDGFRα+ CAFs through PDGFA/PDGFRα signaling. These CAFs engage OSCC cells through two complementary crosstalk pathways: (1) heterotypic E-cadherin/integrin α2β1 adhesion that transmits biomechanical cues to activate YAP signaling and induce epithelial-mesenchymal transition (EMT)-like programs; and (2) tunneling nanotube (TNT)-mediated mitochondrial transfer that enhances oxidative phosphorylation (OXPHOS) and bioenergetic supply in budding cells. Targeted inhibition of TNT-mediated mitochondrial transfer markedly suppresses TB in TBO and xenograft models. Together, our results reveal a mechano-metabolic symbiosis between PDGFRα+/integrin α2+ CAFs and OSCC cells that drives TB and provides actionable targets to block this aggressive metastatic precursor.