An analysis toolbox to explore mesenchymal migration heterogeneity reveals adaptive switching between distinct modes

TOOLS AND RESOURCES An analysis toolbox to explore mesenchymal migration heterogeneity reveals adaptive switching between distinct modes Hamdah Shafqat-Abbasi, Jacob M Kowalewski, Alexa Kiss, Xiaowei Gong, Pablo Hernandez-Varas, Ulrich Berge, Mehrdad Jafari-Mamaghani, John G Lock*†, Staffan Strömblad† Department of Biosciences and Nutrition, Karolinska Institutet, Huddinge, Sweden Abstract Mesenchymal (lamellipodial) migration is heterogeneous, although whether this *For correspondence: john.lock@ ki.se reflects progressive variability or discrete, ’switchable’ migration modalities, remains unclear. We present an analytical toolbox, based on quantitative single-cell imaging data, to interrogate this heterogeneity. Integrating supervised behavioral classification with multivariate analyses of cell motion, membrane dynamics, cell-matrix adhesion status and F-actin organization, this toolbox here enables the detection and characterization of two quantitatively distinct mesenchymal migration modes, termed ’Continuous’ and ’Discontinuous’. Quantitative mode comparisons reveal differences in cell motion, spatiotemporal coordination of membrane protrusion/retraction, and how cells within each mode reorganize with changed cell speed. These modes thus represent distinctive migratory strategies. Additional analyses illuminate the macromolecular- and cellularscale effects of molecular targeting (fibronectin, talin, ROCK), including ’adaptive switching’ between Continuous (favored at high adhesion/full contraction) and Discontinuous (low adhesion/ inhibited contraction) modes. Overall, this analytical toolbox now facilitates the exploration of both spontaneous and adaptive heterogeneity in mesenchymal migration. † DOI: 10.7554/eLife.11384.001 Competing interests: The authors declare that no competing interests exist. Introduction These authors contributed equally to this work Funding: See page 26 Received: 03 September 2015 Accepted: 16 December 2015 Published: 29 January 2016 Reviewing editor: Johanna Ivaska, University of Turku, Finland Copyright Shafqat-Abbasi et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited. Cell migration is a profoundly heterogeneous phenomenon. Indeed, cells can adopt several substantially different migration modalities, including multicellular, amoeboid, and mesenchymal (also termed lamellipodial or lamellipodial-driven) migration, which can all be utilized by a broad range of cell types, as well as lobopodial migration, which has been observed specifically in fibroblasts (Friedl and Wolf, 2010; Sahai, 2005; Petrie and Yamada, 2015; Petrie et al., 2014; Welch, 2015; Friedl and Alexander, 2011). These migration modes represent ’prespecified’ cellular configurations (i.e. cell states) that are favored under particular conditions (Friedl, 2004). Switch-like conversion between these distinct modes is therefore part of the plastic, adaptive/compensatory response of cells to either environmental modulation (Liu et al., 2015; Starke et al., 2014; Ruprecht et al., 2015) or molecular targeting (Sahai et al., 2007; Sanz-Moreno et al., 2008; Somlyo et al., 2003; Wolf, 2003). At a finer scale, heterogeneity is also evident within these migration modes, arising either stochastically or as an adaptive response to changing cues (Geiger et al., 2009; Lämmermann and Sixt, 2009; Lock et al., 2014; Winograd-Katz et al., 2009). Yet, partly due to a lack of adequate quantification, it remains unclear to what extent variation within modes occurs either progressively along a continuum or in a switch-like manner between as yet undefined intramodal subpopulations. Specifically, in the case of amoeboid migration, three discrete sub-modalities Shafqat-Abbasi et al. eLife 2015;5:e11384. DOI: 10.7554/eLife.11384 1 of 29 Tools and resources Cell biology Computational and systems biology eLife digest During an animal’s lifetime, many of its cells will move from one location in the body to another. For example, skin cells can migrate to repair wounds. Prior to migration, cells are usually attached to a scaffold called the extracellular-matrix, which helps to hold them in a particular location within a tissue. Individual cells can move in different ways. During a type of movement called mesenchymal migration, the front end of a cell grows outwards and attaches to a different section of the matrix. The rear of the cell is pulled forward and it detaches from the matrix and retracts, which allows the entire cell to move forward. In contrast, during amoeboid migration, the moving cells are only loosely attached to the matrix and move by gliding. There are large variations in how cells move and they can adopt modes that lie between the two extremes of mesenchymal and amoeboid migration. They can also switch between modes depending on their requirements. Shafqat-Abbasi et al. developed a method to analyse how individual human lung cancer cells move. The method uses software to collect data on cell shape, speed of movement and other features from microscopy images of the migrating cells. The experiments reveal that the cells adopt two distinct migration modes, which Shafqat-Abbasi et al. termed ’Discontinuous’ and ’Continuous’. The majority of cells migrated in the Discontinuous mode, in which cells moved in many different directions. This was caused by a lack of coordination between the outgrowth of the front end of the cell, and the retraction of the back from the matrix. In contrast, in the cells that migrated using the Continuous mode, an outgrowth consistently led to a retraction, which enabled cells to move in one direction. Further experiments revealed that the mode of migration used by the cells is affected by how tightly they are bound to the extracellular-matrix, and the mechanical forces generated inside the cells to drive the movement. Shafqat-Abbasi et al.’s method provides an analytical toolbox that other researchers can use to study the mesenchymal migration of animal cells. DOI: 10.7554/eLife.11384.002 have been observed, and these can co-exist under individual conditions (Welch, 2015; Lämmermann and Sixt, 2009; Yoshida, 2006). By contrast, potentially distinct styles of mesenchymal (lamellipodial) migration, including keratocyte-like (Barnhart et al., 2015; Keren et al., 2008) and fibroblast-like (Abercrombie et al., 1977; Theisen et al., 2012) migration, have been described as arising largely in separate cell types or conditions. Therefore, despite some early suggestions (Lewis et al., 1982), it has remained uncertain to what extent divergent sub-modalities of mesenchymal migration spontaneously emerge in parallel within uniform cell populations and conditions, and whether these modes are truly quantitatively distinct, or instead represent extremes in a broad phenotypic continuum. These questions are imp (...truncated)