
Emphatic visualization of sphingomyelin-rich domains by inter-lipid FRET imaging using fluorescent sphingomyelins
Author(s) -
Masanao Kinoshita,
Hikaru Ano,
Michio Murata,
Kenta Shigetomi,
Junichi Ikenouchi,
Nobuaki Matsumori
Publication year - 2017
Publication title -
scientific reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.24
H-Index - 213
ISSN - 2045-2322
DOI - 10.1038/s41598-017-16361-x
Subject(s) - förster resonance energy transfer , sphingomyelin , fluorescence , biophysics , membrane , fluorescence lifetime imaging microscopy , lipid raft , chemistry , lipid bilayer , vesicle , biology , biochemistry , physics , quantum mechanics
Imaging the distribution of sphingomyelin (SM) in membranes is an important issue in lipid-raft research. Recently we developed novel fluorescent SM analogs that exhibit partition and dynamic behaviors similar to native SM, and succeeded in visualizing lateral domain-segregation between SM-rich liquid-ordered (L o ) and SM-poor liquid-disordered (L d ) domains. However, because the fluorescent contrast between these two domains depends directly on their partition ratio for the fluorescent SMs, domain-separation becomes indeterminate when the distribution difference is not great enough. In this study, we propose the use of inter-lipid Förster resonance energy transfer (FRET) imaging between fluorescent SMs to enhance the contrast of the two domains in cases in which the inter-domain difference in SM distribution is inadequate for conventional monochromic imaging. Our results demonstrate that inter-lipid FRET intensity was significantly higher in the L o domain than in the L d domain, resulting in a clear and distinguishable contrast between the two domains even in poorly phase-separated giant unilamellar vesicles. In addition, we show that inter-lipid FRET imaging is useful for selective visualization of highly condensed assemblies and/or clusters of SM molecules in living cell membranes. Thus, the inter-lipid FRET imaging technique can selectively emphasize the SM-condensed domains in both artificial and biological membranes.