Trans-cis isomerization of lipophilic dyes probing membrane microviscosity in biological membranes and in live cells.
Membrane environment and fluidity can modulate the dynamics and interactions of membrane proteins and can thereby strongly influence the function of cells and organisms in general. In this work, we demonstrate that trans-cis isomerization of lipophilic dyes is a useful parameter to monitor packaging and fluidity of biomembranes. Fluorescence fluctuations, generated by trans-cis isomerization of the thiocarbocyanine dye Merocyanine 540 (MC540), were first analyzed by fluorescence correlation spectroscopy (FCS) in different alcohol solutions. Similar isomerization kinetics of MC540 in lipid vesicles could then also be monitored, and the influence of lipid polarity, membrane curvature, and cholesterol content was investigated. While no influence of membrane curvature and lipid polarity could be observed, a clear decrease in the isomerization rates could be observed with increasing cholesterol contents in the vesicle membranes. Finally, procedures to spatially map photoinduced and thermal isomerization rates on live cells by transient state (TRAST) imaging were established. On the basis of these procedures, MC540 isomerization was studied on live MCF7 cells, and TRAST images of the cells at different temperatures were found to reliably detect differences in the isomerization parameters. Our studies indicate that trans-cis isomerization is a useful parameter for probing membrane dynamics and that the TRAST imaging technique can provide spatial maps of photoinduced isomerization as well as both photoinduced and thermal back-isomerization, resolving differences in local membrane microviscosity in live cells.
Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, Stockholm 106 91, Sweden.
Volodymyr
V
Chmyrov
Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, Stockholm 106 91, Sweden.
Thiemo
T
Spielmann
Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, Stockholm 106 91, Sweden.
Heike
H
Hevekerl
Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, Stockholm 106 91, Sweden.
Jerker
J
Widengren
D002462:Cell Membrane / Q000737:chemistry*; D005260:Female; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000737:chemistry; D007536:Isomerism; D008055:Lipids / Q000737:chemistry*; D061986:MCF-7 Cells; D008954:Models, Biological; D013194:Staining and Labeling / Q000379:methods*; D014783:Viscosity
D005456:Fluorescent Dyes; D007211:Indoles; D008055:Lipids; C098209:VPM chloride