A role of the Ca2+/Mg2+-dependent endonuclease in apoptosis and its inhibition by poly(ADP-ribose) polymerase

Apoptosis is characterized by various cell morphological and biochemical fe atures, one of which is the internucleosomal degradation of genomic DNA. Th e role of the human chromatin-bound Ca2+- and Mg2+-dependent endonuclease ( CME) DNAS1L3 and its inhibition by poly(ADP-ribosyl)ation in the DNA degrad ation that accompanies apoptosis was investigated. The nuclear localization of this endonuclease is the unique feature that distinguishes it from othe r suggested apoptotic nucleases. Purified recombinant DNAS1L3 was shown to cleave nuclear DNA into both high molecular weight and oligonucleosomal fra gments in vitro. Furthermore, exposure of mouse skin fibroblasts expressing DNAS1L3 to inducers of apoptosis resulted in oligonucleosomal DNA fragment ation, an effect not observed in cells not expressing this CME, as well as in a decrease in cell viability greater than that apparent in the control c ells. Recombinant DNAS1L3 was modified by recombinant human poly(ADP-ribose ) polymerase (PARP) in vitro, resulting in a loss of nuclease activity. The DNAS1L3 protein also underwent poly(ADP-ribosyl)ation in transfected mouse skin fibroblasts in response to inducers of apoptosis. The cleavage and in activation of PARP by a caspase-3-like enzyme late in apoptosis were associ ated with a decrease in the extent of DNAS1L3 poly(ADP-ribosyl)ation, which likely releases DNAS1L3 from inhibition and allows it to catalyze the degr adation of genomic DNA.