Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 110, Issue 8, Pages E613-E622Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1216585110
Keywords
SIMS; stable isotope
Categories
Funding
- US Department of Energy [DE-FG02-07ER46471]
- Burroughs Wellcome Fund
- National Institutes of Health [T32 GM070421]
- Laboratory Directed Research and Development funding
- National Institute of Child Health and Human Development
- National Institutes of Health
- National Science Foundation [CHE-1058809]
- Direct For Mathematical & Physical Scien
- Division Of Chemistry [1058809] Funding Source: National Science Foundation
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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 N-15-enriched ions from metabolically labeled N-15-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 N-15-sphingolipid microdomains with mean diameters of similar to 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.
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