Periodic Reporting for period 3 - STARNEL (supracellular contractility of myofibroblasts in gut homeostasis and cancer invasion) Reporting period: 2021-10-01 to 2023-03-31 Summary of the context and overall objectives of the project There has been tremendous progress in understanding the importance of the microenvironment and its chemical signals for homeostasis of stem cell niche in the intestine and for tumor invasion and metastasis formation in many different tissues. However, the way stromal cells such as myofibroblasts or cancer-associated fibroblasts (CAFs) use mechanical forces to shape the extracellular matrix and consequently dictate the response of epithelial cells remains unexplored at the single-cell level mainly due to limited imaging tools. Here we propose a multi-disciplinary approach, at the interface of cancer cell biology and physics, aimed to understand how myofibroblasts contractility influences epithelial cell functions in physiological (homeostasis) and pathological (cancer) conditions using the gut as a model. Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far During tumor progression, CAFs accumulate in tumors and produce an excessive extracellular matrix (ECM), forming a capsule that enwraps cancer cells. This capsule is a barrier that restricts tumor growth leading to the buildup of intratumoral pressure. Combining genetic and physical manipulations in vivo with microfabrication and force measurements in vitro, we found that the CAFs capsule is not a passive barrier but instead actively compresses cancer cells using actomyosin contractility. Cancer cells mechanosense CAF compression, resulting in an altered localization of the transcriptional regulator YAP. Abrogation of CAFs contractility in vivo leads to the dissipation of compressive forces and impairment of capsule formation. By mapping CAF force patterns in 3D, we show that compression is a CAF-intrinsic property independent of cancer cell growth. Supracellular coordination of CAFs is achieved through fibronectin cables that serve as scaffolds allowing force transmission. This study unveils that the contractile capsule actively compresses cancer cells, modulates their mechanical signaling, and reorganizes tumor morphology. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) There has been tremendous progress in understanding the importance of the microenvironment and its chemical signals for homeostasis of stem cell niche in the intestine and for tumor invasion and metastasis formation in many different tissues. However, the way the stromal cells such as myofibroblasts or CAFs use mechanical forces to shape the ECM and consequently dictate the response of epithelial cells remains unexplored at the single-cell level, mainly due to limited imaging tools. Using a multi-disciplinary approach at the interphase between cancer cell biology and physics, we expect to unravel the fundamental mechanisms and roles of myofibroblasts supracellular contractility in intestinal homeostasis and cancer progression. We believe that our work will shed light on the pathophysiological consequences of cell contractility and will generate novel ideas for anti-cancer therapies by targeting its microenvironment. CAFs compress cancer cells
