Complex regulation of tumor necrosis factor mRNA turnover in lipopolysaccharide-activated macrophages

Biochim Biophys Acta. 1991 Aug 27;1090(1):22-8. doi: 10.1016/0167-4781(91)90032-h.

Abstract

The turnover of tumor necrosis factor (TNF) mRNA in permanently transfected macrophages of the RAW 264.7 cell line was studied directly (by Northern blot analysis using a probe specific for TNF) and indirectly (through studies of the turnover of various reporter mRNAs, either containing or lacking the TNF 3' untranslated region (UTR)). The TNF mRNA was found to be very unstable in RAW 264.7 cells. Instability appeared to result from two distinguishable nucleolytic processes. The major degradative process involved was not specific for the TNF 3' UTR of reporter mRNAs, and was inhibited by actinomycin D pretreatment. It appeared to be expressed constitutively, in that cell activation by lipopolysaccharide (LPS) did not modify message stability. When cells were treated with actinomycin D, a minor nucleolytic activity was 'uncovered'. This minor activity was noted to increase with time following LPS activation. It also exhibited specificity, in that reporter mRNAs bearing the 3' UTR of TNF were more susceptible to degradation in the presence of actinomycin D than were constructs lacking the 3' UTR of TNF. Thus, TNF mRNA turnover appears complex, and depends upon at least two separable degradative pathways. The TNF 3' UTR apparently contributes only modestly to the instability of this mRNA under normal conditions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Northern
  • Cell Line
  • Dactinomycin / pharmacology
  • Gene Expression Regulation / physiology*
  • Kinetics
  • Lipopolysaccharides / pharmacology
  • Macrophage Activation / physiology
  • Macrophages / drug effects
  • Macrophages / metabolism*
  • Mice
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism*
  • Tumor Necrosis Factor-alpha / genetics*

Substances

  • Lipopolysaccharides
  • RNA, Messenger
  • Tumor Necrosis Factor-alpha
  • Dactinomycin