Depletion of RIPK3 or MLKL blocks TNF-driven necroptosis and switches towards a delayed RIPK1 kinase-dependent apoptosis

Cell Death Dis. 2014 Jan 16;5(1):e1004. doi: 10.1038/cddis.2013.531.

Abstract

In human cells, the RIPK1-RIPK3-MLKL-PGAM5-Drp1 axis drives tumor necrosis factor (TNF)-induced necroptosis through mitochondrial fission, but whether this pathway is conserved among mammals is not known. To answer this question, we analyzed the presence and functionality of the reported necroptotic axis in mice. As in humans, knockdown of receptor-interacting kinase-3 (RIPK3) or mixed lineage kinase domain like (MLKL) blocks TNF-induced necroptosis in L929 fibrosarcoma cells. However, repression of either of these proteins did not protect the cells from death, but instead induced a switch from TNF-induced necroptosis to receptor-interacting kinase-1 (RIPK1) kinase-dependent apoptosis. In addition, although mitochondrial fission also occurs during TNF-induced necroptosis in L929 cells, we found that knockdown of phosphoglycerate mutase 5 (PGAM5) and dynamin 1 like protein (Drp1) did not markedly protect the cells from TNF-induced necroptosis. Depletion of Pink1, a reported interactor of both PGAM5 and Drp1, did not affect TNF-induced necroptosis. These results indicate that in these murine cells mitochondrial fission and Pink1 dependent processes, including Pink-Parkin dependent mitophagy, apparently do not promote necroptosis. Our data demonstrate that the core components of the necrosome (RIPK1, RIPK3 and MLKL) are crucial to induce TNF-dependent necroptosis both in human and in mouse cells, but the associated mechanisms may differ between the two species or cell types.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis*
  • Dynamins / genetics
  • Dynamins / metabolism
  • Humans
  • Mice
  • Necrosis
  • Phosphoprotein Phosphatases
  • Phosphoric Monoester Hydrolases / genetics
  • Phosphoric Monoester Hydrolases / metabolism
  • Protein Kinases / genetics
  • Protein Kinases / metabolism*
  • Receptor-Interacting Protein Serine-Threonine Kinases / genetics
  • Receptor-Interacting Protein Serine-Threonine Kinases / metabolism*
  • Tumor Necrosis Factors / genetics
  • Tumor Necrosis Factors / metabolism*

Substances

  • Tumor Necrosis Factors
  • MLKL protein, mouse
  • Protein Kinases
  • Receptor-Interacting Protein Serine-Threonine Kinases
  • Ripk1 protein, mouse
  • Ripk3 protein, mouse
  • PGAM5 protein, mouse
  • Phosphoprotein Phosphatases
  • Phosphoric Monoester Hydrolases
  • Dnm1l protein, mouse
  • Dynamins