Mouse granzyme A induces a novel death with writhing morphology that is mechanistically distinct from granzyme B-induced apoptosis

Cell Death Differ. 2013 Sep;20(9):1183-93. doi: 10.1038/cdd.2013.59. Epub 2013 Jun 7.

Abstract

Human and mouse granzyme (Gzm)B both induce target cell apoptosis in concert with pore-forming perforin (Pfp); however the mechanisms by which other Gzms induce non-apoptotic death remain controversial and poorly characterised. We used timelapse microscopy to document, quantitatively and in real time, the death of target cells exposed to primary natural killer (NK) cells from mice deficient in key Gzms. We found that in the vast majority of cases, NK cells from wild-type mice induced classic apoptosis. However, NK cells from syngeneic Gzm B-deficient mice induced a novel form of cell death characterised by slower kinetics and a pronounced, writhing, 'worm-like' morphology. Dying cells initially contracted but did not undergo membrane blebbing, and annexin-V staining was delayed until the onset of secondary necrosis. As it is different from any cell death process previously reported, we tentatively termed this cell death 'athetosis'. Two independent lines of evidence showed this alternate form of death was due to Gzm A: first, cell death was revealed in the absence of Gzm B, but was completely lost when the NK cells were deficient in both Gzm A and B; second, the athetotic morphology was precisely reproduced when recombinant mouse Gzm A was delivered by an otherwise innocuous dose of recombinant Pfp. Gzm A-mediated athetosis did not require caspase activation, early mitochondrial disruption or generation of reactive oxygen species, but did require an intact actin cytoskeleton and was abolished by latrunculin B and mycalolide B. This work defines an authentic role for mouse Gzm A in granule-induced cell death by cytotoxic lymphocytes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actin Cytoskeleton
  • Animals
  • Apoptosis / drug effects*
  • Apoptosis / immunology
  • Bridged Bicyclo Compounds, Heterocyclic / pharmacology
  • Cell Line, Tumor
  • Granzymes / deficiency
  • Granzymes / genetics
  • Granzymes / metabolism*
  • HeLa Cells
  • Humans
  • Killer Cells, Natural / cytology
  • Killer Cells, Natural / immunology*
  • Marine Toxins
  • Mice
  • Mice, Inbred C57BL
  • Microscopy, Confocal
  • Oxazoles / pharmacology
  • Perforin / metabolism*
  • Thiazolidines / pharmacology
  • Time-Lapse Imaging

Substances

  • Bridged Bicyclo Compounds, Heterocyclic
  • Marine Toxins
  • Oxazoles
  • Thiazolidines
  • mycalolide B
  • Perforin
  • Granzymes
  • latrunculin B