TGF-β2 enhances nanoscale cortex stiffness via condensation of cytoskeleton-focal adhesion plaque

Physical spatiotemporal characteristics of cellular cortex dominate cell functions and even determine cell fate. The cellular cortex is able to reorganize to a dynamic steady status with changed stiffnesses once stimulated, and thus alter the physiological and pathological activities of almost all types of cells. TGF-β2, a potent pleiotropic growth factor, plays important roles in cartilage development, endochondral ossification, and cartilage diseases. However, it is not yet known whether TGF-β2 would alter the physical spatiotemporal characteristics of the cell cortex such as cortex stiffness, thereby affecting the function of chondrocytes. In this study, we investigated the influence of TGF-β2 on cellular cortex stiffness of chondrocytes and the underlying mechanism. We firstly detected TGF-β2-induced changes in cytoskeleton and focal adhesion plaque, which were closely related to cellular cortex stiffness. We then characterized the landscape of nanoscale cortex stiffness in individual chondrocytes induced by TGF-β2 via atomic force microscopy. By using inhibitors, latrunculin A and blebbistatin, we verified the importance of cytoskeleton-focal adhesion plaque axis on cellular cortex stiffness of chondrocytes induced by TGF-β2. We finally elucidated that TGF-β2 enhanced the phosphorylation of Smad3 and facilitated the nuclear accumulation of p-Smad3. The p-Smad3 aggregated in the nuclei enhanced the cytoskeleton and focal adhesion plaque at transcriptional level, thereby mediating changes in cell cortex stiffness. Taken together, these results provide an understanding about the role of TGF-β2 on physical spatiotemporal properties of cell cortex in chondrocytes, and might provide cues for interpretation of cartilage development and interventions to cartilage diseases.