Phosphorylation of interleukin-6 at serine54: an early event in the secretory pathway in human fibroblasts.
The cytokine interleukin-6 (IL-6) is the major phosphoprotein secreted by human fibroblasts induced with interleukin-1 alpha (IL-1 alpha). We have determined that Ser54 is the predominant site of phosphorylation on the fibroblast-derived IL-6 polypeptide; the amino acid motif surrounding this site is reminiscent of the target site for the Golgi-associated protein (casein) kinase. It has been shown earlier that the IL-6 polypeptide follows the classical secretory pathway where N-linked glycosylation is detectable within the first 15 minutes of labeling with [35S]-methionine and O-linked glycosylation occurs between 15-30 minutes after the start of polypeptide synthesis. Pulse-chase experiments using [32P]-orthophosphate or [35S]-methionine as tracers indicated that phosphorylation of IL-6 occurred prior to its O-glycosylation suggesting that the de novo synthesized IL-6 polypeptide is rapidly, perhaps even cotranslationally, phosphorylated at an intravesicular site (in the endoplasmic reticulum and/or Golgi). When IL-1 alpha-induced fibroblasts were exposed to cycloheximide there was enhancement of the net de novo synthesis and secretion of IL-6 as followed by [35S]-methionine labeling ("superinduction") but the secreted cytokine was no longer phosphorylated as monitored by [32P] labeling. Thus, phosphorylation of the IL-6 polypeptide is not an obligatory requirement for secretion of this cytokine. Furthermore, IL-6 phosphorylation is inhibited by cycloheximide through a mechanism different from the drug's effects on polypeptide synthesis per se.
May
L T
LT
Department of Microbiology & Immunology, New York Medical College, Valhalla 10595.
Sehgal
P B
PB
D000595:Amino Acid Sequence; D003513:Cycloheximide / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D015850:Interleukin-6 / Q000378:metabolism*; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism
D007375:Interleukin-1; D015850:Interleukin-6; D010750:Phosphoproteins; D010768:Phosphoserine; D003513:Cycloheximide