The resistance of circulating tumor cells to large fluid shear stress in hematogenous dissemination by reducing mechanosensitivity via nuclear mechanosensing mediated myosin cytoplasmic redistribution

Abstract

Tumor cells metastasize to distant organs mainly through hematogenous dissemination, where circulating tumor cells (CTCs) experience varying levels of fluid shear stress (FSS) in capillaries, veins, and arteries. Although FSS is one critical rate-limiting factor of cell survival, there are always a small subpopulation of CTCs persisting within the vasculature, which may eventually generate metastatic tumors that contribute to the majority of cancer-related death. It is well known that living cells that adhere to solid substrates can perceive mechanical cues through mechanotransduction. However, how CTCs in suspension respond to varying levels of FSS remains unclear. Importantly, limited knowledge of how CTCs mechanically adapt to FSS impedes our understanding of CTCs’ survival during hematogenous dissemination. In this study, we found that the apoptosis of suspended breast tumor cells (SBTCs) showed weaker increase rate under high FSS compared to low FSS range, suggesting that SBTCs may exhibit reduced mechanosensitivity under high FSS, which could serve as a protective mechanoadaptation mechanism for their survival. Mechanistically, the phosphorylated myosin light chain (P-MLC), representative of myosin II activation, was predominantly localized in the cortex under low FSS and re-distributed to the cytoplasm under high FSS. Similar phenomena were observed in SBTCs that were attached to Poly-L-Lysine (PLL) coated glass, which could mimic the suspension status. Further, we defined the ratio of the resultant nuclear strain and the applied FSS as force transmission efficiency (FTE) from the cell membrane to nucleus, which was relatively high under low FSS but decreased under high FSS in both suspended and PLL-adhered tumor cells. Inhibition of cytoplasmic but not cortical P-MLC pharmacologically and specifically restored the FTE under high FSS and enhanced shear-induced DNA damage and cell apoptosis. To elucidate the mechanism underlying P-MLC subcellular relocalization, we discovered that Lamin A/C-mediated nuclear mechanosensing increased nuclear envelop tension and triggered calcium release from endoplasmic reticulum, which further activated cortical and cytoplasmic P-MLC through myosin light chain kinase (MLCK) and Rho associated protein kinase (ROCK) under low and high FSS, respectively. Targeting ROCK but not MLCK rescued shear-induced decease in FTE and increased the shear-induced apoptosis of SBTCs both in vitro and in vivo. In addition, high cytoplasmic P-MLC was also found in cancer stem cells (CSCs) and blood cells that held the survival advantage under FSS. Pharmacologic inhibition of ROCK significantly increased the shear-induced cell apoptosis. Collectively, our results unveiled a previously unappreciated role of nuclear mechanosensing mediated cytoplasmic myosin II activation in regulating force transmission and mechanosensitivity of SBTCs, by which CTCs mechanically adapted to large FSS, providing new insights into the survival of CTCs during hematogenous metastasis.