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Endonuclease activation causing genomic degradation is a pervasive hallmark of apoptosis and a suggested precipitating or commitment step in the suicidal process. Directly applied endonuclease activity has produced apoptotic-like effects in isolated nuclei, but not yet shown as an initiating apoptogen in whole cells. Mechanistically genomic damage inflicted by a variety of DNA-damaging agents is also known to produce mitotic catastrophe condensations characterizing cell cycle derangement. Morphological and molecular similarities between apoptosis and mitotic catastrophe have been noted, but their conjoint expressions from directly applied endonuclease activity has also not been shown. We show here micrococcal nuclease (MNase) initiating apoptosis in human Chang liver cells which expressed both apoptotic and mitotic catastrophe condensations. Genomic profiling showed (a) the two stage apoptotic sequence of large (50 kb) and small (200 bp) fragment cleavage demonstrated by pulse field and normal gel electrophoresis, respectively; (b) the sub-G1 ;apoptotic peak' with shrunken cells from flow cytometric evaluation of PI-DNA binding and laser forward scattering, (c) 3' OH termini typical of apoptotic DNA fragments labelled by terminal deoxynucleotidyl transferase (TdT)-mediated fluorescence tagging especially in the shrunken cells, and (d) positive comet assay of the apoptotic genome. Nuclear shrinkage evaluated by confocal image analysis was consistent with the apoptotic response, as was Zn2+ ion sensitivity, an established inhibitor of apoptotic expression. Endonuclease activity per se is apoptogenetic and mechanistically convergent with the mitotic catastrophe pathway in the proliferative cycle.

cell death and differentiation

Micrococcal nuclease (endonuclease) digestion causes apoptosis and mitotic catastrophe with interphase chromosome condensation in human Chang liver cells