Genome digestion is a dispensable consequence of physiological cell death mediated by cytotoxic T lymphocytes.

Abstract

We examined virally transformed murine fibroblast clones as targets for cytotoxic T lymphocyte (CTL)-triggered lysis and genome digestion. Strikingly, while all clones were essentially equivalent in the ability to be lysed, one clone, SV3T3-B2.1, failed to exhibit genome digestion associated with CTL attack. Other aspects of the physiological cell death process, including loss of adhesion and nuclear envelope breakdown (lamin phosphorylation and solubilization), were not altered in this clone. The absence of genome digestion associated with CTL-induced cell death correlated with the absence of endodeoxyribonuclease activity in the nuclei of that clone. Characterization of the activity affected identifies a calcium-dependent, DNase I-like endonuclease of approximately 40 kDa, normally present constitutively in all cell nuclei, as the enzyme responsible for genome digestion associated with CTL-mediated cell death. These observations indicate that neither genome digestion per se nor its consequences [such as activation of poly(ADP-ribose) polymerase] are essential for cell death resulting from the triggering of this cell suicide process.Extensive genome digestion has been taken as the hallmark of physiological cell death (for examples, see references 2, 10, 11, 31, and 39). Cell deaths occur selectively in many tissue types throughout development and in homeostasis. These physiological deaths appear to follow an orderly process of internal cellular disintegration that is, generally, cell autonomous and dependent on macromolecular synthesis. The process includes nuclear envelope breakdown, cytoskeletal reorganization and plasma membrane blebbing, and loss of adhesion; some of these morphological characteristics have been termed apoptosis. Associated with breakdown of the nuclear envelope is digestion of genomic DNA. Dying lymphoid cells undergo extensive genome digestion, typically generating a ladder of double-stranded oligonucleosomal DNA fragments visualized by gel electrophoresis. In contrast, dying nonlymphoid cells often exhibit much less genome digestion; denaturing sucrose gradient centrifugation can reveal the infrequent single-stranded nicks (21,28). Pathological cell deaths, including those resulting from membrane damage (produced by antibody and complement or cytotoxic pore-forming molecules, for example), are not associated with orderly cellular disintcgration and prelytic genome digestion (reviewed in reference 57).Target cell death mediated by cytotoxic T lymphocytes (CTL) is a unique case of physiological cell death. Target cell death is not cell autonomous and does not require target cell macromolecular synthesis (33,47). Nonetheless, the active participation of the targeted cell appears to be required (59), and in many morphological and biochemical respects, the cell death process is indistinguishable from cell autonomous suicides. Indeed, the concept of internal disintegration, with prelytic nuclear events including genome digestion preced-* Corresponding author.ing the loss of plasma membrane int...

Resistance of Ehrlich tumor cells to apoptosis can be due to accumulation of heat shock proteins

“…Both hyperthermia and ATP depletion were found to induce protein aggregation, and suppression of the aggregation in stressed cells with the elevated HSP content [9,16] may be responsible for the protection from these apoptosis-inducing exposures. Suggested suppression of actin aggregation by HSPs [21] is of special importance since G-actin (but not F-actin) is a potent inhibitor of DNase I which can be involved in apoptotic DNA fragmentation [22][23][24].…”

Apoptotic DNA fragmentation is detected by a semi-quantitative ligation-mediated PCR of blunt DNA ends

“…Little is known about the endonuclease(s) responsible for ladder production besides its dependence on pH and the presence of Ca 2+ or Mg 2+ ions in mammals (Cohen and Duke, 1984;Arends et al, 1990). Several candidate nucleases have been proposed, including DNase I (Ucker et al, 1992;Peitsch et al, 1993), DNase II (Barry and Eastman, 1993) and Nuc 18 (Caron-Leslie et al, 1991); hence, identifying enzymes which produce DNA fragments with blunt, 5' phosphorylated ends may help to discern the nucleases that have a physiological role. We note that the only endonuclease activity shown to generate these types of DNA ends in vivo is the V(D)J recombinase encoded in part by RAG-1 and RAG-2 (Schlissel et al, 1993) which could conceivably participate in ladder production in the thymus, and perhaps the CNS where RAG-1 is expressed (Chun et al, 1991).…”

Herbimycin A accelerates the induction of apoptosis following etoposide treatment or γ-irradiation of bcr/abl-positive leukaemia cells

“…However, the percentage of TUNEL-positive cells generated in the presence of the drug combination was approximately half the percentage of cells showing nuclear condensation under the same conditions. Others have also observed that DNA degradation is not always detected in cells undergoing apoptosis as de®ned by morphological criteria (Tomei et al, 1993;Ucker et al, 1992).…”