Ubiquitin-like protein MNSFβ covalently binds to Bcl-G and enhances lipopolysaccharide/interferon γ-induced apoptosis in macrophages

FEBS J. 2013 Mar;280(5):1281-93. doi: 10.1111/febs.12120. Epub 2013 Feb 11.


Monoclonal non-specific suppressor factor β (MNSFβ) is a ubiquitously expressed member of the ubiquitin-like family that is involved in various biological functions. Previous studies have demonstrated that MNSFβ covalently binds to intracellular pro-apoptotic protein Bcl-G and regulates the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) cascade in the mouse macrophage cell line Raw264.7. In this study, we demonstrate that MNSFβ promotes lipopolysaccharide (LPS)/interferon γ (IFNγ)-induced apoptosis of Raw264.7 macrophages. In Raw264.7 cells treated with MNSFβ small interfering RNA (siRNA), LPS/IFNγ- or NO donor S-nitrosoglutathione-induced apoptosis was inhibited. siRNA-mediated knockdown of MNSFβ did not affect inducible nitric-oxide synthase (iNOS) expression in LPS/IFNγ-stimulated Raw264.7 cells. Conversely, co-transfection with MNSFβ and Bcl-G greatly enhanced LPS/IFNγ- induced apoptosis in Raw264.7 cells, accompanied by increased expression of p53 and decreased Cox-2 activity. Unlike co-transfection with wild-type MNSFβ, co-transfection of a mutant MNSFβ (G74A) and Bcl-G did not result in enhancement of LPS/IFNγ-induced apoptosis. Co-over-expression of MNSFβ and Bcl-G reduced S-nitrosoglutathione-induced ERK1/2 phosphorylation. Furthermore, electrophoretic mobility shift assay experiments revealed that MNSFβ down-regulates the ERK/activator protein 1 (AP-1) signaling cascade which leads to Cox-2 activation. We also observed that MNSFβ-Bcl-G promotes LPS/IFNγ-induced apoptosis of mouse peritoneal macrophages, together with a decrease in Cox-2 expression. Taken together, our data indicate an apoptosis-enhancing effect of MNSFβ-Bcl-G is due in part to down-regulation of Cox-2 activation in macrophages.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis / drug effects*
  • Blotting, Western
  • Cells, Cultured
  • Cyclooxygenase 2 / genetics
  • Cyclooxygenase 2 / metabolism
  • Immunoprecipitation
  • Interferon-gamma / pharmacology*
  • Lipopolysaccharides / pharmacology*
  • Macrophages, Peritoneal / drug effects
  • Macrophages, Peritoneal / metabolism
  • Macrophages, Peritoneal / pathology*
  • Mice
  • Mice, Inbred BALB C
  • Nitric Oxide / metabolism
  • Nitric Oxide Synthase Type II / genetics
  • Nitric Oxide Synthase Type II / metabolism
  • Phosphorylation
  • Protein Binding
  • Proto-Oncogene Proteins c-bcl-2 / genetics
  • Proto-Oncogene Proteins c-bcl-2 / metabolism*
  • RNA, Messenger / genetics
  • RNA, Small Interfering / genetics
  • Real-Time Polymerase Chain Reaction
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Suppressor Factors, Immunologic / antagonists & inhibitors
  • Suppressor Factors, Immunologic / genetics
  • Suppressor Factors, Immunologic / metabolism*


  • BCL2L14 protein, mouse
  • Lipopolysaccharides
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Messenger
  • RNA, Small Interfering
  • Recombinant Proteins
  • Suppressor Factors, Immunologic
  • monoclonal nonspecific suppressor factor
  • Nitric Oxide
  • Interferon-gamma
  • Nitric Oxide Synthase Type II
  • Ptgs2 protein, mouse
  • Cyclooxygenase 2