Convection-Induced Biased Distribution of Actin Probes in Live Cells |
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Authors: | Sawako Yamashiro Daisuke Taniguchi Soichiro Tanaka Tai Kiuchi Dimitrios Vavylonis Naoki Watanabe |
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Institution: | 1. Laboratory of Single-Molecule Cell Biology, Kyoto University Graduate School of Medicine, Kyoto Japan;2. Department of Pharmacology, Kyoto University Graduate School of Medicine, Kyoto Japan;3. Laboratory of Single-Molecule Cell Biology, Tohoku University Graduate School of Life Sciences, Sendai, Miyagi, Japan;4. Department of Physics, Lehigh University, Bethlehem, Pennsylvania |
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Abstract: | Fluorescent markers that bind endogenous target proteins are frequently employed for quantitative live-cell imaging. To visualize the actin cytoskeleton in live cells, several actin-binding probes have been widely used. Among them, Lifeact is the most popular probe with ideal properties, including fast exchangeable binding kinetics. Because of its fast kinetics, Lifeact is generally believed to distribute evenly throughout cellular actin structures. In this study, however, we demonstrate misdistribution of Lifeact toward the rear of lamellipodia where actin filaments continuously move inward along the retrograde flow. Similarly, phalloidin showed biased misdistribution toward the rear of lamellipodia in live cells. We show evidence of convection-induced misdistribution of actin probes by both experimental data and physical models. Our findings warn about the potential error arising from the use of target-binding probes in quantitative live imaging. |
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Keywords: | Corresponding author |
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