Raft-based sphingomyelin interactions revealed by new fluorescent sphingomyelin analogs

Masanao Kinoshita, Kenichi G.N. Suzuki, Nobuaki Matsumori, Misa Takada, Hikaru Ano, Kenichi Morigaki, Mitsuhiro Abe, Asami Makino, Toshihide Kobayashi, Koichiro M. Hirosawa, Takahiro K. Fujiwara, Akihiro Kusumi, Michio Murata

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43 Citations (Scopus)

Abstract

Sphingomyelin (SM) has been proposed to form cholesterol-dependent raft domains and sphingolipid domains in the plasma membrane (PM). How SM contributes to the formation and function of these domains remains unknown, primarily because of the scarcity of suitable fluorescent SM analogs. We developed new fluorescent SM analogs by conjugating a hydrophilic fluorophore to the SM choline headgroup without eliminating its positive charge, via a hydrophilic nonaethylene glycol linker. The new analogs behaved similarly to the native SM in terms of their partitioning behaviors in artificial liquid order-disorder phase-separated membranes and detergent-resistant PM preparations. Single fluorescent molecule tracking in the live-cell PM revealed that they indirectly interact with each other in cholesterol- and sphingosine backbone-dependent manners, and that, for ~10-50 ms, they undergo transient colocalization-codiffusion with a glycosylphosphatidylinositol (GPI)-anchored protein, CD59 (in monomers, transient-dimer rafts, and clusters), in CD59-oligomer size-, cholesterol-, and GPI anchoring-dependent manners. These results suggest that SM continually and rapidly exchanges between CD59-associated raft domains and the bulk PM.

Original languageEnglish
Pages (from-to)1183-1204
Number of pages22
JournalJournal of Cell Biology
Volume216
Issue number4
DOIs
Publication statusPublished - Apr 1 2017

All Science Journal Classification (ASJC) codes

  • Cell Biology

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    Kinoshita, M., Suzuki, K. G. N., Matsumori, N., Takada, M., Ano, H., Morigaki, K., Abe, M., Makino, A., Kobayashi, T., Hirosawa, K. M., Fujiwara, T. K., Kusumi, A., & Murata, M. (2017). Raft-based sphingomyelin interactions revealed by new fluorescent sphingomyelin analogs. Journal of Cell Biology, 216(4), 1183-1204. https://doi.org/10.1083/jcb.201607086