Development and Cancer

Mechanical forces, which are ubiquitous in nature, are well known to drive tissue formation and impact cellular function. These forces are crucial in regulating cell morphogenesis, migration and even adhesion to the extracellular matrix. Mechanical forces therefore impact a wide variety of biological processes, from cell proliferation and differentiation to tissue mass homeostasis and metastasis – all processes towards which our research is focused.

A human foreskin fibroblast, plated on fibronection, stained for the actin cytoskeleton (green) and imaged on a spinning disk microscope. Image captured by Tee Yee Han of the Bershadsky lab.

Making Contacts

The physical forces necessary for cells to attach themselves to the substrate and to one another is key to cellular movement. The points of contact act as mechanical sensors that provide the cell with information about its environment and so determine its behavior. Integrin-based adhesion complexes (focal adhesions), which form a major point of cell-matrix contact, are a key research focus at the MBI. These complexes are closely associated with the actin cytoskeleton and so connect the intra- and extracellular environments. They are able to recognize a variety of stimuli, not only in the form of biochemical factors, but also physical and geometrical attributes. We use micro- and nano-fabrication techniques, such as the production of micropillars, to measure and test the effect of such attributes as elasticity, dimensionality and ligand spacing on the cell.

In cancer cells the activity of these mechanical sensors is disrupted, which likely accounts for the cell’s difficulty in adhering to substrates and consequently their greater mobility. Mechanical signals are therefore suggested to have a role in tumor progression, making efforts to understand the process of mechanotransduction highly relevant to cancer.

A mouse embryonic fibroblast of the NIH3T3 cell line plated on collagen, stained for actin (green), paxillin (magenta) and p130CAS (cyan) and captured on a spinning disk microscope. This image, highlighting stress fibers and cell-matrix adhesions, was captured by Alvin Guo of the Sawada lab.

Different matrices, different fates

The differentiation pathway chosen by a cell depends on matrix signals, of both a biochemical and biophysical nature. Alterations in the rigidity of the extracellular matrix can therefore result in alterations in cellular function. Changes in cellular function ultimately alter tissue morphogenesis, which is dependent on the coordinated spatial and temporal generation of cellular forces. The use of both cell lines and Drosophila allow us to study the effects of spatial and geometric constraints of the microenvironment on cell and tissue dynamics. In order to advance our understanding of collective cell migration, we measure the forces exerted when tissues translocate across a surface, establish the locations of these forces and further aim to identify the cellular machinery responsible for generating these forces.

A differentiating rat adrenal medulla cell of the PC12 cell line expressing lysosomal marker LAMP1 (lysosomal-associated membrane protein 1) (red) imaged on a spinning disk microscope. Image captured by Aarthi Ravichandran of the Low lab.

Signals and Sensors

Signal transduction molecules, both regulators and effectors, play important roles in controlling cell growth, death, differentiation, migration and tissue development. The dynamic forces and geometrical influences of the environment that impinge on cells and tissues are translated through an extensive network of GTPases. This family of proteins forms a key point of study in our endeavour to understand the process of mechanotransduction.

Specific force receptors, termed mechanosensors, are directly modulated by mechanical stimuli and are able to initiate signaling cascades. With the exception of mechanosensitive channels, few have been identified at a molecular level. We propose that ion channel-independent mechanosensors exist, which deform in response to physical forces and are working towards identifying and characterizing these sensors. In order to investigate signal transduction processes at the cell membrane, we combine aspects of cellular biophysics, physical chemistry and materials science.

A mouse embryonic fibroblast stained for the actin cytoskeleton (red) and nucleus (blue) and captured on a spinning disk microscope. This image of stress fibers was captured by Alvin Guo of the Sawada lab.

Tools of the Trade

The development of sophisticated tools such as FRET (Fluorescence resonance energy transfer) and super-resolution microscopy have allowed us to study how proteins are assembled into nanoscale machinery and how this machinery interacts with the environment. Through the creation of 3D microenvironments to study different aspects of cell-cell adhesion, we aim to shed more light on these critical points of contact.

Selected Publications

Min Wu, Bo Huang, Morven Graham, Andrea Raimondi, John E. Heuser, Xiaowei Zhuang & Pietro De Camilli. Coupling between clathrin-dependent endocytic budding and F-BAR-dependent tubulation in a cell-free system. Nature Cell Biology, 12, 902–908(2010)

P. Kanchanawong, G. Shtengel, A. Pasapera-Limon, E. Ramko, M.W. Davidson, H.F. Hess, C. M. Waterman. Nanoscale Architecutre of Integrin-based Adhesions. Nature, 468(7323): 580-4(2010)

Toyama, Y., Peralta, X.G., Wells, A.R., Kiehart, D.P., and Edwards, G.S. ‘Apoptotic force and tissue dynamics during Drosophila embryogenesis’ Science 321, 1683 (2008).

Zhou, Y.T, Chew, L.L, Lin, S.C, Low, B.C. The BNIP-2 and Cdc42GAP homology (BCH) domain of p50RhoGAP/Cdc42GAP sequesters RhoA from inactivation by the adjacent GTPase-activating protein domain. Mol Biol Cell., 2010 21(18):3232-46(2010)

Masha Prager-Khoutorsky, Alexandra Lichtenstein, Ramaswamy Krishnan, Kavitha Rajendran, Avi Mayo, Zvi Kam, Benjamin Geiger* & Alexander D. Bershadsky*. Fibroblast polarization is a matrix-rigidity-dependent process controlled by focal adhesion mechanosensing. Advanced Online Publication at Nature Cell Biology, DOI (Digital Object Identifier) 10.1038/ncb2370 *corresponding authors.

Sawada, Y., M. Tamada, B.J. Dubin-Thaler, O. Cherniavskaya, R. Sakai, S. Tanaka & M.P. Sheetz. Force Sensing by Mechanical Extension of the Src Family Kinase Substrate p130Cas. Cell, 127:1015-26(2006)

Wong CH, Chan H, Ho CY, Lai SK, Chan KS, Koh CG* & Hoi-Yeung Li*. Apoptotic histone modification inhibits nuclear transport by regulating RCC1. Nature Cell Biology, 11: 36-45(2009). *corresponding authors.

Wong, S.Y., Chiam, K.H., Lim,C.T. & Matsudaira,P. Computational model of cell positioning, directed and collective migration in the intestinal crypt epithelium. J.R. Soc.Interface 7, S351-363 (2010)

Ryu S and P. Matsudaira. Unsteady Motion, Finite Reynolds Number and Wall Effects on Vorticella convallaria Contribute Contraction Force Greater than the Stokes Drag. Biophys. J., 98:2574-81(2010)

Luke H. Chao, Margaret M. Stratton, Il-Hyung Lee, Oren S. Rosenberg, Joshua Levitz, Daniel J. Mandell, Tanja Kortemme, Jay T. Groves, Howard Schulman & John Kuriyan. A Mechanism for Tunable Autoinhibition in the Structure of a Human Ca2+/Calmodulin- Dependent Kinase II Holoenzyme. Cell, 146(5), 732-745(2011)