Spreading of Nuclear Degradation in Pinus thunbergii Parl. Cuttings under Water Stress

The killing of suspension-cultured tobacco cells and stem cellsof Japanese black pine (Pinus thunbergii Parl.) by drying, autoclaving,hypertonic treatment or freezing caused the disappearance ordeformation of nuclei, as monitored by staining with 4',6-diamidino-2-phenylindole(DAPI). This observation suggests that disappearance or deformationof the nucleus can be taken as an index of cell death. Usingthis index, we examined the spreading of the cell death in water-stressedcuttings of Japanese black pine. Various portions of the cuttingswere examined over time and DAPI-stained nuclei were seen todisappear more rapidly in the upper than in the lower portionsof water-stressed cuttings. In every portion, undeformed, DAPI-stainednuclei disappeared sequentially from the cortex and the pith,from the periderm, from the xylem and finally from the cambium,indicating that cell death caused by water stress spreads fromthe cortex and the pith to the periderm, then to the xylem,and finally to the cambium. Water conduction was ex amined byimmersing the cut ends of cuttings in a red solution of acidfuchsin. Under water stress, water conduction was reduced andthe height of the upper limit of the sucked solution above thecut end was reduced. Just below this upper limit, water conductionwas inhibited in some regions of the xylem. However, undeformed,DAPI-stained nuclei were distributed homogeneously over bothconductive and non-conductive regions of the xylem, indicatingthat there is no direct relationship between cell death andwater conductivity of the xylem. Undeformed, DAPI-stained nucleidisappeared in regions around the upper limit of suction ofthe dye, and the sequence of disappearance was again from thecortex and the pith, from the periderm, from the xylem and,finally, from the cambium. (Received January 16, 1992; Accepted July 14, 1992)