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Participation of de novo sphingolipid biosynthesis in the regulation of autophagy in Kdo 2 ‐Lipid A stimulated RAW264.7 macrophages
Author(s) -
Sims Kacee,
Haynes Christopher,
Kelly Samuel,
Allegood Jeremy,
Wang Elaine,
Momin Amin,
Leipelt Martina,
Reichart Donna,
Glass Christopher,
Sullards M. Cameron,
Merrill Alfred
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.906.4
Subject(s) - sphingolipid , microbiology and biotechnology , ceramide , autophagy , sphingomyelin , biology , golgi apparatus , lactosylceramide , biosynthesis , sphingosine , lipid signaling , biochemistry , vacuole , cell culture , cell , chemistry , receptor , cytoplasm , glycolipid , membrane , enzyme , genetics , apoptosis
Activation of RAW264.7 cells with a lipopolysaccharide specific for the TLR4 receptor, Kdo 2 ‐Lipid A (KLA), causes a large increase in cellular sphingolipids‐‐from 1.5 to 2.6 × 109 molecules per cell in 24 h, based on the sum of subspecies analyzed by “lipidomic” mass spectrometry. Thus, this study asked: What is the cause of this increase, and is there a cell function connected with it? The sphingolipids arise primarily from de novo biosynthesis; with the exception of ceramide, which is also produced from pre‐existing sources. Nonetheless, the activated RAW264.7 cells have a higher number of sphingolipids per cell because KLA inhibits cell division, thus, the cells are larger and contain increased numbers of membrane vacuoles termed autophagosomes. Indeed, de novo biosynthesis of sphingolipids performs an essential structural and/or signaling function in autophagy because autophagosome formation was eliminated by ISP1 in KLA stimulated RAW264.7 cells; furthermore, an anti‐ceramide antibody co‐localizes with autophagosomes in activated RAW264.7 cells versus the Golgi in unstimulated or ISP1 inhibited cells. These findings establish that KLA induces profound changes in sphingolipid metabolism and content in this macrophage‐like cell line, apparently to produce sphingolipids that are necessary for formation of autophagosomes, which are thought to play important roles in the mechanisms of innate immunity.