Chromatin extrusion in resting encystment of Colpoda cucullus: a possible involvement of apoptosis-like nuclear death

The extrusion of macronuclear chromatin is a remarkable characteristic during encystment in Colpoda, but the biological significance of this phenomenon has not been fully elucidated. Here we demonstrate that chromatin extrusion occurs with high frequency when encystment was induced by increasing Ca(2+) in growing cells in various stages of the cell cycle. The Feulgen-DNA reaction revealed that vegetatively growing cells have more macronuclear DNA than cells in the stationary phase, suggesting an association of macronuclear DNA content with the execution of chromatin extrusion. Using 4',6-diamidino-2-phenylindole (DAPI), we found that the size of the macronuclear extrusion body was reduced with time and eventually disappeared approximately 24h after encystment induction. In addition, oligonucleosome-sized DNA cleavage was confirmed to occur concomitant with the size reduction, suggesting that the extrusion body is selectively degraded, while the normal macronucleus remains alive. Combined use of acridine orange and Hoechst 33342 demonstrated that the extruded body was increasingly acidified before final resorption. These features are reminiscent of the nuclear degradation process in conjugating Tetrahymena, and therefore we conclude that chromatin extrusion in Colpoda might occur to adjust the macronuclear DNA content prior to encystment. In this way, it is similar to the apoptotic-like nuclear death that occurs during the conjugation of other ciliates.