Biology subjects

Vosatka, K. W.

Publications and source records attributed to Vosatka, K. W..

2 recordsLinked to original sources

bioRxiv2021

A novel Fiji/ImageJ-based macro identifies the limitations of F-actin markers in studying the fast amoeboid migration of confined cells

When confined, cells have recently been shown to undergo a phenotypic switch to what has been termed, fast amoeboid (leader bleb-based) migration. However, as this is a nascent area of research, few tools are available for the rapid analysis of cell behavior. Here, we demonstrate that a novel Fiji/ImageJ-based plugin, Analyze_Blebs, can be used to quickly obtain cell migration parameters and morphometrics from time lapse images. As validation, we show that Analyze_Blebs can detect significant differences in cell migration and morphometrics, such as the largest bleb size, upon introducing different live markers of F-actin, including F-tractin and LifeAct tagged with green and red fluorescent proteins. We also demonstrate, using flow cytometry, that live markers increase total levels of F-actin. Furthermore, that F-tractin increases cell stiffness, which was found to correlate with a decrease in migration, thus reaffirming the importance of cell mechanics as a determinant of Leader Bleb-Based Migration (LBBM). Highlight summaryA new plugin, Analyze_Blebs, enables the rapid analysis of cell migration and morphometrics of fast amoeboid cells. Morphometrics combined with cell stiffness are found to be predictive of confined, fast amoeboid (leader bleb-based), migration.

cell biology↗

bioRxiv2020

Emerin regulation of nuclear stiffness is required for the amoeboid migration of cancer cells in confining environments

When metastasizing, tumor cells must traverse environments with diverse physicochemical properties. Recently, the cell nucleus has emerged as a major regulator of the transition from mesenchymal to fast amoeboid (leader bleb-based) migration. Here, in melanoma cells, we demonstrate that increasing nuclear stiffness through elevating Lamin A, inhibits fast amoeboid migration. Importantly, nuclei may respond to force through stiffening. A key factor in this process is the inner nuclear membrane (INM) protein, emerin. Accordingly, we determined the role of emerin in regulating fast amoeboid migration. Strikingly, we found that both the up- and down-regulation of emerin results in an inhibition of fast amoeboid migration. However, when key Src phosphorylation sites were removed, up-regulation of emerin no longer inhibited fast amoeboid migration. Interestingly, in confined cells, Src activity was low, as measured by a Src biosensor. Thus, the fast amoeboid migration of melanoma cells depends on the precise calibration of emerin activity. Summary StatementIn mechanically constrictive microenvironments, amoeboid migrating melanoma cells require emerin for the precise calibration of nuclear stiffness.

cell biology↗