Loss of giant obscurins alters breast epithelial cell mechanosensing of matrix stiffness
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Kimberly M. Stroka1, Bin Sheng Wong2,3,4, Marey Shriver5, Jude M. Phillip2,3,4, Denis Wirtz2,3,4, Aikaterini Kontrogianni-Konstantopoulos5,6 and Konstantinos Konstantopoulos2,3,4
1Fischell Department of Bioengineering, University of Maryland, College Park, MD, 20742, USA
2Johns Hopkins Institute for NanoBioTechnology, The Johns Hopkins University, Baltimore, MD, 21218, USA
3Johns Hopkins Physical Sciences-Oncology Center, The Johns Hopkins University, Baltimore, MD, 21218, USA
4Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, MD, 21218, USA
5University of Maryland School of Medicine, Department of Biochemistry and Molecular Biology, Baltimore, MD, 21201, USA
6University of Maryland School of Medicine, Marlene and Stewart Greenebaum National Cancer Institute Cancer Center, Baltimore, MD, 21201, USA
Kimberly M. Stroka, email: [email protected]
Konstantinos Konstantopoulos, email: [email protected]
Keywords: matrix stiffness, mechanosensitivity, cell migration, obscurin, RhoA
Received: June 07, 2016 Accepted: July 20, 2016 Published: August 01, 2016
Obscurins are a family of RhoGEF-containing proteins with tumor and metastasis suppressing roles in breast epithelium. Downregulation of giant obscurins in normal breast epithelial cells leads to reduced levels of active RhoA and of its downstream effectors. Herein, we elucidate how depletion of giant obscurins affects the response of breast epithelial cells to changes in the mechanical properties of the microenvironment. We find that knockdown of obscurins increases cell morphodynamics, migration speed, and diffusivity on polyacrylamide gels of ≥ 1 kPa, presumably by decreasing focal adhesion area and density as well as cell traction forces. Depletion of obscurins also increases cell mechanosensitivity on soft (0.4–4 kPa) surfaces. Similar to downregulation of obscurins, pharmacological inhibition of Rho kinase in breast epithelial cells increases migration and morphodynamics, suggesting that suppression of Rho kinase activity following obscurin knockdown can account for alterations in morphodynamics and migration. In contrast, inhibition of myosin light chain kinase reduces morphodynamics and migration, suggesting that temporal changes in cell shape are required for efficient migration. Collectively, downregulation of giant obscurins facilitates cell migration through heterogeneous microenvironments of varying stiffness by altering cell mechanobiology.
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