The extracellular matrix (ECM) regulates carcinogenesis by interacting with cancer cells via cell surface receptors. Discoidin Domain Receptor 2 (DDR2) is a collagen-activated receptor implicated in cell survival, growth, and differentiation. Dysregulated DDR2 expression has been identified in various cancer types, making it as a promising therapeutic target. Additionally, cancer cells exhibit mechanosensing abilities, detecting changes in ECM stiffness, which is particularly important for carcinogenesis given the observed ECM stiffening in numerous cancer types. Despite these, whether collagen-activated DDR2 signaling and ECM stiffness-induced mechanosensing exert similar effects on cancer cell behavior and whether they operate through analogous mechanisms remain elusive. To address these questions, we performed bulk RNA sequencing (RNA-seq) on human SH-SY5Y neuroblastoma cells cultured on collagen-coated substrates. Our results show that DDR2 downregulation induces significant changes in the cell transcriptome, with changes in expression of 15% of the genome, specifically affecting the genes associated with cell division and differentiation. We validated the RNA-seq results by showing that DDR2 knockdown redirects the cell fate from proliferation to senescence. Like DDR2 knockdown, increasing substrate stiffness diminishes cell proliferation. Surprisingly, RNA-seq indicates that substrate stiffness has no detectable effect on the transcriptome. Furthermore, DDR2 knockdown influences cellular responses to substrate stiffness changes, highlighting a crosstalk between these two ECM-induced signaling pathways. Based on our results, we propose that the ECM could activate DDR2 signaling and mechanosensing in cancer cells to orchestrate their cell fate through distinct mechanisms, with or without involving gene expression, thus providing novel mechanistic insights into cancer progression. © 2024 The Authors. FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.