2D protrusion but not motility predicts growth factor–induced cancer cell migration in 3D collagen

Meyer, Aaron S.; Hughes, Shannon K.; Kay, Jennifer E.; Castillo, Amalchi; Wells, Alan; Gertler, Frank B.; Lauffenburger, Douglas A.

doi:10.1083/jcb.201201003

Cited by 89 publications

(84 citation statements)

References 40 publications

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“…The trends for directional protrusion extension are strikingly similar to the trends for the direction of cell migration (Figs. 4G and 5D), consistent with the observation that protrusion formation is a marker for 3D motility of MDA-MB-231 cells (33).…”

Section: Discussionsupporting

confidence: 75%

“…Actin polymerization and protrusion formation are required for cell migration in 3D ECMs, and the number of protrusions per cell in 2D scales with motility of MDA-MB 231 cells in 3D (33). Exposing cells to 4.6 μm/s flow for 4 h caused a decrease in protrusion formation, but cells extended more protrusions in the upstream direction (Fig.…”

Section: Paxillin Is Required For Upstream Actin Polarization and Promentioning

confidence: 99%

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Mechanotransduction of fluid stresses governs 3D cell migration

Polacheck

German

Mammoto

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

225

215

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Solid tumors are characterized by high interstitial fluid pressure, which drives fluid efflux from the tumor core. Tumor-associated interstitial flow (IF) at a rate of ∼3 μm/s has been shown to induce cell migration in the upstream direction (rheotaxis). However, the molecular biophysical mechanism that underlies upstream cell polarization and rheotaxis remains unclear. We developed a microfluidic platform to investigate the effects of IF fluid stresses imparted on cells embedded within a collagen type I hydrogel, and we demonstrate that IF stresses result in a transcellular gradient in β1-integrin activation with vinculin, focal adhesion kinase (FAK), FAK PY397 , F actin, and paxillindependent protrusion formation localizing to the upstream side of the cell, where matrix adhesions are under maximum tension. This previously unknown mechanism is the result of a force balance between fluid drag on the cell and matrix adhesion tension and is therefore a fundamental, but previously unknown, stimulus for directing cell movement within porous extracellular matrix.mechanobiology | breast cancer | metastasis I ntegrins and associated focal adhesion (FA) proteins form a tension-sensitive mechanical link between the extracellular matrix (ECM) and the cytoskeleton, and serve as key components in the signaling cascade by which cells transduce mechanical signals into biological responses (mechanotransduction) (1, 2). Contractile stresses generated by the cell are balanced by tractions at cell-substrate adhesions, and the FA protein vinculin accumulates at regions of high substrate stress (3, 4). The FA protein paxillin colocalizes with vinculin (4) and mediates β1-integrin FA turnover through interaction with FA kinase (FAK) (5). The FAK-paxillin signaling axis recruits vinculin to β1 integrins at regions of high matrix adhesion tension (6), and paxillin-a key mechanosensor (7)-mediates protrusion formation at regions of high stress on 2D substrates (8), and FAK-paxillin-vinculin signaling is required for mechanosensing and durotaxis (9).The tumor microenvironment imparts mechanical and chemical signals on tumor and stromal cells (10), and advanced breast carcinomas are characterized by high interstitial fluid pressure (11), an indicator of poor prognosis (12). This elevated fluid pressure drives interstitial flow (IF) and alters chemical transport within the tumor (13), and IF influences tumor cell migration through the generation of autocrine chemokine gradients (14). Equally important, although not as well understood, is the physical drag imparted on the ECM and constitutive cells (15) by IF, which is analogous to the FA-activating shear stresses generated on endothelial cells by hemodynamic forces (16). With endothelial cells, shear stress can be the dominant mechanical stimulus that induces FAK activation and cytoskeletal remodeling; however, for cells embedded within a porous matrix scaffold, the ratio of the force due to the pressure drop across the cell to the total shear force is inversely proportional to hydrogel p...

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Section: Discussionsupporting

confidence: 75%

Section: Paxillin Is Required For Upstream Actin Polarization and Promentioning

confidence: 99%

Mechanotransduction of fluid stresses governs 3D cell migration

Polacheck

German

Mammoto

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

225

215

View full text Add to dashboard Cite

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“…While RhoC depleted cells show 2D locomotion with no change in overall cell velocity, protrusion formation is dramatically inhibited. Recent results from Lauffenburger lab (Meyer et al, 2012) show that acute regulation of protrusion dynamics in 2D (but not other parameters like speed or persistence) reflects long-term 3D migration response and this correlates with the invasive behavior of tumor cells. These results are consistent with our findings and suggest that protrusion formation is a key indicator of migratory behavior of invasive tumor cells in the tumor microenvironment and highlights the importance of elucidating the mechanisms of protrusion formation to understand complex processes like tumor metastasis.…”

Section: Discussionmentioning

confidence: 99%

Spatial regulation of RhoC activity defines protrusion formation in migrating cells

Bravo‐Cordero

Sharma

Roh-Johnson

et al. 2013

Journal of Cell Science

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SummaryProtrusion formation is the first step that precedes cell movement of motile cells. Spatial control of actin polymerization is necessary to achieve directional protrusion during cell migration. Here we show that the spatial coordinators p190RhoGEF and p190RhoGAP regulate actin polymerization during leading edge protrusions by regulating the actin barbed end distribution and amplitude. The distribution of RhoC activity and proper balance of cofilin activation achieved by p190RhoGEF and p190RhoGAP determines the direction of final protrusive activity. These findings provide a new insight into the dynamic plasticity in the amplitude and distribution of barbed ends, which can be modulated by fine-tuning RhoC activity by upstream GEFs and GAPs for directed cell motility.

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“…The ability of cells to generate membrane protrusions plays an important role in numerous biological activities, particularly in cell migration and invasion (37)(38)(39)(40), which are mainly mediated by protrusions in the form of filopodia and lamellipodia (41). BloCPrinting provides a rapid and high-throughput method to characterize cell protrusions, including protrusion percentage and extension rate, and cell length, which are challenging to achieve using existing methods.…”

Section: Gap Junction Intercellular Communication In Cell Pairs With mentioning

confidence: 99%

Block-Cell-Printing for live single-cell printing

Zhang¹,

Chou

Xia

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

127

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A unique live-cell printing technique, termed "Block-Cell-Printing" (BloC-Printing), allows for convenient, precise, multiplexed, and high-throughput printing of functional single-cell arrays. Adapted from woodblock printing techniques, the approach employs microfluidic arrays of hook-shaped traps to hold cells at designated positions and directly transfer the anchored cells onto various substrates. BloC-Printing has a minimum turnaround time of 0.5 h, a maximum resolution of 5 μm, close to 100% cell viability, the ability to handle multiple cell types, and efficiently construct protrusion-connected single-cell arrays. The approach enables the large-scale formation of heterotypic cell pairs with controlled morphology and allows for material transport through gap junction intercellular communication. When six types of breast cancer cells are allowed to extend membrane protrusions in the BloC-Printing device for 3 h, multiple biophysical characteristics of cells-including the protrusion percentage, extension rate, and cell length-are easily quantified and found to correlate well with their migration levels. In light of this discovery, BloC-Printing may serve as a rapid and high-throughput cell protrusion characterization tool to measure the invasion and migration capability of cancer cells. Furthermore, primary neurons are also compatible with BloC-Printing.cell array | cell communication | protrusion profiling | neuron patterning C urrent high-throughput screening of cell function and heterogeneity and in vitro cell-cell communication studies requires routine generation of large-scale single-cell arrays with high precision and efficiency, single-cell resolution, multiple cell types, and maintenance of cell viability and function (1, 2). Several approaches have been designed for this purpose, e.g., inkjet cell printing (3-6), surface engineering (7-15), and physical constraints (16-23). However, finding a method that completely satisfies the above requirements remains a challenge. Potentially useful and convenient tools may be available by adapting traditional printing tools to cell printing. In particular, woodblock printing is an efficient and convenient technology that revolutionized the printing world more than 1,800 y ago and was extended to microcontact molecular printing ∼20 y ago (24). However, application of the block-printing concept to cells has never been achieved. The main challenges are (i) inking cells to their molds with precision and maintaining viability, (ii) evenly and gently applying and transferring the cells to a substrate and successfully lifting off the mold without detaching the cells, and (iii) maintaining cell functions after printing.We report here the development and testing of a technology called "Block-Cell-Printing" (BloC-Printing), which involves directly inking cells to a predesigned mold and then transferring the cells to a substrate. We overcome the challenges described above by flow patterning, instead of pressing, the ink objects, as in woodblock or microcontact printing. By ...

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2D protrusion but not motility predicts growth factor–induced cancer cell migration in 3D collagen

Abstract: In breast cancer cells, growth factor stimulation of membrane protrusion was a better predictor of 3D migration than 2D motility, cognate receptor expression, or receptor activation.

Cited by 89 publications

References 40 publications

Mechanotransduction of fluid stresses governs 3D cell migration

Mechanotransduction of fluid stresses governs 3D cell migration

Spatial regulation of RhoC activity defines protrusion formation in migrating cells

Block-Cell-Printing for live single-cell printing

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