Processing of newly synthesized cachectin/tumor necrosis factor in endotoxin-stimulated macrophages.
The biosynthesis and processing of cachetin/tumor necrosis-factor (TNF) were examined in the murine macrophage-like cell line RAW 264.7. Lipopolysaccharide-stimulated cells secreted both glycosylated and nonglycosylated 17-kilodalton (kDa) mature cachectin/TNF into the culture medium. Secreted cachectin/TNF was derived from membrane-associated precursors that were precipitated by polyclonal antisera raised against either the mature protein or synthetic peptide fragments of the 79 amino acid cachectin/TNF prohormone sequence. About half of the precursors were N-glycosylated, apparently cotranslationally. The cachectin/TNF precursors were then proteolytically cleaved to release soluble mature cytokine into the medium, while the membrane-bound 14-kDa presequence remained cell associated. During the period of LPS stimulation, the amount of macrophage cell surface cachectin/TNF remained at a low level, suggesting that both nonglycosylated and glycosylated precursors of cachectin/TNF are efficiently cleaved by these cells. These findings suggest the presence of a unique mechanism for the secretion of cachectin/TNF.
Jue
D M
DM
Laboratory of Medical Biochemistry, Rockefeller University, New York, New York 10021.
Sherry
B
B
Luedke
C
C
Manogue
K R
KR
Cerami
A
A
D000818:Animals; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004731:Endotoxins / Q000494:pharmacology; D006031:Glycosylation; D007739:L Cells / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D011498:Protein Precursors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000378:metabolism
D004731:Endotoxins; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D011498:Protein Precursors; D014409:Tumor Necrosis Factor-alpha