Chromatin Collapse during Caspase-dependent Apoptotic Cell Death Requires DNA Fragmentation Factor, 40-kDa Subunit-/Caspase-activated Deoxyribonuclease-mediated 3′-OH Single-strand DNA Breaks

Abstract

Background: Apoptotic nuclear morphology can occur independently of DFF40/CAD-mediated DNA fragmentation. Results: DFF40/CAD induces 3Ј-OH single-strand DNA nicks/breaks and nuclear collapse during caspase-dependent apoptosis. Conclusion: Caspase-dependent apoptotic nuclear collapse is prompted by DFF40/CAD-mediated single-strand DNA damage. Significance: The knowledge of how apoptotic nuclear collapse occurs should be relevant to understand the final steps of cell demise and its influence on the cellular environment.

Caspase-activated DNase Is Necessary and Sufficient for Oligonucleosomal DNA Breakdown, but Not for Chromatin Disassembly during Caspase-dependent Apoptosis of LN-18 Glioblastoma Cells

“…In fact, this is the first report on a cellular model in which the compaction of the chromatin is independent of the enzymatic action of DFF40/CAD. Our finding appears to be discordant with previous data establishing that the alteration of DFF40/ CAD function abolishes both chromatin compaction and nuclear apoptotic body formation (10,12,33,36,37). An alternative explanation might be that DFF40/CAD-mediated apoptotic signaling is necessary but not sufficient to induce the complete chromatin reorganization during apoptosis, and other DFF40/CAD-independent intracellular pathways are involved.…”

“…Indeed, DFF40/CAD knockdown avoids stage II nuclear morphology (revealing stage I nuclear morphology) and oligonucleosomal DNA degradation in cells proficient to display any of the apoptotic hallmarks (33). In the same sense, the overexpression of this endonuclease is enough to facilitate the apoptotic hallmarks in those cells that are defective in displaying them (33). In LN-18 cells, as expected, the overexpression of DFF40/CAD provides the ability to degrade the chromatin into oligonucleosomal length fragments.…”

“…In all cases, cells need functional DFF40/CAD to display both hallmarks after caspase activation. Indeed, DFF40/CAD knockdown avoids stage II nuclear morphology (revealing stage I nuclear morphology) and oligonucleosomal DNA degradation in cells proficient to display any of the apoptotic hallmarks (33). In the same sense, the overexpression of this endonuclease is enough to facilitate the apoptotic hallmarks in those cells that are defective in displaying them (33).…”

“…However, DFF40/CAD is still required. In this regard, we have recently reported that the nuclear collapse and disassembly observed during caspase-dependent cell death in SK-N-AS cells, which fail to degrade their DNA into oligonucleosome-sized fragments depend on DFF40/CAD catalytic activity (33). LN-18 cells although undergoing caspase-dependent cell death, are unable to degrade their genomic content into oligonucleosome-sized fragments, irrespectively of the drug employed.…”

AChE for DNA degradation

“…However, whether AChE-S is a true apoptotic DNase in cellula needs to be further confirmed. In this sense, and unfortunately, the authors only performed TUNEL assay in AChE-Soverexpressing cells and, as recently described, TUNEL-positive cells do not necessarily undergo DNA laddering [8]. Therefore, further efforts are required to address this key issue.…”