Chromatin breakdown during necrosis by serum Dnase1 and the plasminogen system.
OBJECTIVE
Dnase1-deficient mice with the 129 x C57BL/6 genetic background develop symptoms of systemic lupus erythematosus, such as high titers of antinuclear autoantibodies directed against nucleosomes. In this study we analyzed a potential molecular pathomechanism leading to this autoimmunity, by exploring the influence of extracellular Dnase1 present in serum on the breakdown of chromatin in necrotic cells in vitro.


METHODS
Human breast adenocarcinoma cells (MCF-7) and other cell lines were subjected to necrosis induced by hydrogen peroxide, streptolysin O, or freeze-thawing. Subsequently, the influence of sera from Dnase1-deficient and wild-type mice as well as the influence of purified enzymes present in the culture medium on the process of necrotic chromatin breakdown was investigated.


RESULTS
Necrotic chromatin breakdown resembled apoptotic DNA laddering and was catalyzed by serum Dnase1 in conjunction with plasmin. During necrosis, Dnase1 and plasminogen penetrated the cell and accumulated in the cytoplasm and nucleus. Plasminogen bound to the cytoskeleton and nuclear structures, was activated to plasmin by either tissue-type or urokinase-type plasminogen activator, and degraded histone H1, thereby facilitating internucleosomal DNA cleavage by Dnase1.


CONCLUSIONS
Our results suggest that serum Dnase1 in cooperation with the plasminogen system guarantees a fast and effective breakdown of chromatin during necrosis by the combined cleavage of DNA as well as of DNA binding proteins. The failure of such a clearance mechanism might lead to antinuclear autoimmunity similar to that observed in the Dnase1-deficient mouse.
Napirei
Markus
M
Abteilung für Anatomie und Embryologie, Ruhr-Universität Bochum, Bochum, Germany. markus.napirei@ruhr-unibochem.de
Wulf
Swantje
S
Mannherz
Hans Georg
HG
D000230:Adenocarcinoma; D000818:Animals; D000974:Antibodies, Antinuclear / Q000502:physiology*; D015551:Autoimmunity / Q000502:physiology*; D001943:Breast Neoplasms; D002843:Chromatin / Q000201:enzymology* / Q000276:immunology / Q000473:pathology; D004247:DNA / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000097:blood*; D005110:Extracellular Space / Q000378:metabolism; D018922:HL-60 Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains; D041681:NIH 3T3 Cells; D009336:Necrosis; D010958:Plasminogen / Q000502:physiology*
D000974:Antibodies, Antinuclear; D002843:Chromatin; D006657:Histones; D010958:Plasminogen; D004247:DNA; C085693:DNASE1 protein, human; D003850:Deoxyribonuclease I