5OH DNA Breaks in Apoptosis and Their Labeling by Topoisomerase-Based Approach
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Recently, the concept of apoptotic cell elimination was expanded and programed cell death is no longer viewed as an individual
cellular event. The complete description of the apoptotic process now includes two phases: the self-driven cell disassembly
and the externally-controlled elimination of apoptotic cell corpses by phagocytizing cells. The second, phagocytic phase is
essential, highly conserved, and is even more important than the internal phase of cell disassembly. This is because it ensures
the complete degradation of the dying cell’s DNA, preventing the release of pathological, viral and tumor DNA, and self-immunization.
In different cells and species from mammals to flies, a single conserved enzyme – DNase II is responsible for the elimination
of cellular DNA in the second “mopping up” phase of apoptosis. Here, we present an assay for the selective detection of the
phagocytic phase of apoptosis. The technology capitalizes on the fact that phagocytic DNase II produces identifiable signature
DNA breaks, which can be labeled by vaccinia topoisomerase. The assay permits labeling of the previously underestimated phase
of apoptotic execution and is a useful tool in the apoptosis detection arsenal.