Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts

Jessica F. Frisz, Kaiyan Lou, Haley A. Klitzing, William P. Hanafin, Vladimir Lizunov, Robert L. Wilson, Kevin J. Carpenter, Raehyun Kim, Ian D. Hutcheon, Joshua Zimmerberg, Peter K. Weber, Mary L. Kraft

Research output: Contribution to journalArticle

Abstract

Sphingolipids play important roles in plasma membrane structure and cell signaling. However, their lateral distribution in the plasma membrane is poorly understood. Here we quantitatively analyzed the sphingolipid organization on the entire dorsal surface of intact cells by mapping the distribution of 15N-enriched ions from metabolically labeled 15N- sphingolipids in the plasmamembrane, using highresolution imagingmass spectrometry.Many types of control experiments (internal, positive, negative, and fixation temperature), along with parallel experiments involvingthe imaging offluorescent sphingolipids-both in living cells and during fixation of living cells-exclude potential artifacts. Micrometer-scale sphingolipid patches consisting of numerous 15N-sphingolipid microdomains with mean diameters of ∼200 nm are always present in the plasma membrane. Depletion of 30% of the cellular cholesterol did not eliminate the sphingolipid domains, but did reduce their abundance and longrange organization in the plasma membrane. In contrast, disruption of the cytoskeleton eliminated the sphingolipid domains. These results indicate that these sphingolipid assemblages are not lipid rafts and are instead a distinctly different type of sphingolipid-enriched plasma membrane domain that depends upon cortical actin.

Original languageEnglish (US)
Pages (from-to)E613-E622
JournalProceedings of the National Academy of Sciences of the United States of America
Volume110
Issue number8
DOIs
StatePublished - Feb 19 2013

Keywords

  • SIMS
  • Stable isotope

ASJC Scopus subject areas

  • General

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    Frisz, J. F., Lou, K., Klitzing, H. A., Hanafin, W. P., Lizunov, V., Wilson, R. L., Carpenter, K. J., Kim, R., Hutcheon, I. D., Zimmerberg, J., Weber, P. K., & Kraft, M. L. (2013). Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts. Proceedings of the National Academy of Sciences of the United States of America, 110(8), E613-E622. https://doi.org/10.1073/pnas.1216585110