Loss of splicing factor ASF/SF2 induces G2 cell cycle arrest and apoptosis, but inhibits internucleosomal DNA fragmentation

Abstract

ASF/SF2 is an SR protein splicing factor that participates in constitutive and alternative pre-mRNA splicing and is essential for cell viability. Using a genetically modified chicken B-cell line, DT40-ASF, we now show that ASF/SF2 inactivation results in a G2-phase cell cycle arrest and subsequent programmed cell death. However, although several hallmarks of apoptosis are apparent, internucleosomal DNA fragmentation was not detected. Furthermore, inactivation of ASF/SF2 also blocks DNA fragmentation normally induced by a variety of apoptotic stimuli. Notably, mRNA encoding the inhibitor of caspase-activated DNase-L (ICAD-L), which acts as an inhibitor as well as a chaperone of caspase-activated DNase (CAD), decreased in abundance, whereas the level of mRNA encoding ICAD-S, which has only inhibitory activity, increased upon ASF/SF2 depletion. Strikingly, expression of appropriate levels of exogenous human ICAD-L restored apoptotic DNA laddering in ASF/SF2-depleted cells. These results not only indicate that loss of an SR protein splicing factor can induce cell cycle arrest and apoptosis, but also illustrate the important role of ICAD and its regulation by alternative splicing in the process of apoptotic DNA fragmentation.