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Title: Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts [High-Resolution Chemical Imaging of Sphingolipid Distribution in the Plasma Membrane]

Abstract

Sphingolipids play important roles in plasma membrane structure and cell signaling. Yet, 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 plasma membrane using high-resolution imaging mass spectrometry. Many types of control experiments (internal, positive, negative, and fixation temperature), along with parallel experiments involving the imaging of fluorescent sphingolipids$-$both in living cells and during fixation of living cells$-$exclude potential artifacts. Micrometer-scale sphingolipid patches consisting of numerous 15Nsphingolipid 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 long range 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.

Authors:
 [1];  [1];  [1];  [1];  [2];  [1];  [3];  [1];  [3];  [2];  [3];  [1]
  1. Univ. of Illinois, Urbana, IL (United States). School of Chemical Sciences
  2. National Inst. of Health, Bethesda, MD (United States). Program in Physical Biology, Eunice Kennedy Shriver National Inst. of Child Health and Human Development
  3. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States). Glenn T. Seaborg Inst.
Publication Date:
Research Org.:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org.:
USDOE; National Institutes of Health (NIH); National Science Foundation (NSF)
OSTI Identifier:
1396208
Report Number(s):
LLNL-JRNL-463256
Journal ID: ISSN 0027-8424
Grant/Contract Number:  
AC52-07NA27344; FG02-07ER4647; CHE-1058809
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Volume: 110; Journal Issue: 8; Journal ID: ISSN 0027-8424
Publisher:
National Academy of Sciences, Washington, DC (United States)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL AND ANALYTICAL CHEMISTRY; SIMS; stable isotope

Citation Formats

Frisz, Jessica F., Lou, Kaiyan, Klitzing, Haley A., Hanafin, William P., Lizunov, Vladimir, Wilson, Robert L., Carpenter, Kevin J., Kim, Raehyun, Hutcheon, Ian D., Zimmerberg, Joshua, Weber, Peter K., and Kraft, Mary L. Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts [High-Resolution Chemical Imaging of Sphingolipid Distribution in the Plasma Membrane]. United States: N. p., 2013. Web. doi:10.1073/pnas.1216585110.
Frisz, Jessica F., Lou, Kaiyan, Klitzing, Haley A., Hanafin, William P., Lizunov, Vladimir, Wilson, Robert L., Carpenter, Kevin J., Kim, Raehyun, Hutcheon, Ian D., Zimmerberg, Joshua, Weber, Peter K., & Kraft, Mary L. Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts [High-Resolution Chemical Imaging of Sphingolipid Distribution in the Plasma Membrane]. United States. doi:10.1073/pnas.1216585110.
Frisz, Jessica F., Lou, Kaiyan, Klitzing, Haley A., Hanafin, William P., Lizunov, Vladimir, Wilson, Robert L., Carpenter, Kevin J., Kim, Raehyun, Hutcheon, Ian D., Zimmerberg, Joshua, Weber, Peter K., and Kraft, Mary L. Mon . "Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts [High-Resolution Chemical Imaging of Sphingolipid Distribution in the Plasma Membrane]". United States. doi:10.1073/pnas.1216585110. https://www.osti.gov/servlets/purl/1396208.
@article{osti_1396208,
title = {Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts [High-Resolution Chemical Imaging of Sphingolipid Distribution in the Plasma Membrane]},
author = {Frisz, Jessica F. and Lou, Kaiyan and Klitzing, Haley A. and Hanafin, William P. and Lizunov, Vladimir and Wilson, Robert L. and Carpenter, Kevin J. and Kim, Raehyun and Hutcheon, Ian D. and Zimmerberg, Joshua and Weber, Peter K. and Kraft, Mary L.},
abstractNote = {Sphingolipids play important roles in plasma membrane structure and cell signaling. Yet, 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 plasma membrane using high-resolution imaging mass spectrometry. Many types of control experiments (internal, positive, negative, and fixation temperature), along with parallel experiments involving the imaging of fluorescent sphingolipids$-$both in living cells and during fixation of living cells$-$exclude potential artifacts. Micrometer-scale sphingolipid patches consisting of numerous 15Nsphingolipid 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 long range 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.},
doi = {10.1073/pnas.1216585110},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
issn = {0027-8424},
number = 8,
volume = 110,
place = {United States},
year = {2013},
month = {1}
}

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