Dynamic Changes in Distribution of Lignin and Hemicelluloses in Cell Walls During Differentiation of Secondary Xylem in Eucommia ulmoides

The dynamic changes in the distribution of lignin and hemicelluloses (xylans and xyloglucans) in cell walls during the differentiation of secondary xylem in Eucommia ulmoides Oliv. were studied by means of ultraviolet light microscopy and transmission electron microscopy combined with immunogold labelling. In the cambial zone and cell expansion zone, xyloglucans were localized both in the tangential and radial walls, but no xylans or lignin were found in these regions. With the formation of secondary wall S-1 layer, lignin occurred in the cell corners and middle lamella, while xylans appeared in S-1 layer, and xyloglucans were localized in the primary walls and middle lamella. In pace with the formation of secondary wall S-2 and S-3 layer, lignification extended to S-1, S-2 and S-3 layer in sequence, showing a patchy style of lignin deposition. Concurrently, xylans distributed in the whole secondary walls and xyloglucans, on the other hand, still localized in the primary walls and middle lamella. The results indicated that along with the formation and lignification of the secondary wall, great changes had taken place in the cell walls. Different parts of cell walls, such as cell corners, middle lamella, primary walls and various layers of secondary walls, had different kinds of hemicelluloses, which formed various cell wall architecture combined with lignin and other cell wall components.

Ultracytochemical Localization of ATPase During the Secondary Xylem Differentiation and Dedifferentiation in Eucommia ulmoides Trunk

The ultracytochemical localization of ATPase in the secondary xylem cells during their differentiation and dedifferentiation in the girdled Eucommia ulmoides Oliv. was carried out using a lead phosphate precipitation technique. Throughout the differentiation, which is a typical programmed cell death (PCD) process, ATPase deposits increased in the nucleus but decreased and progressively disappeared in the cell organelles. At the same time, the distribution of ATPase increased in the inner face of the cell wall and pits with cytoplasmic degeneration. The results demonstrated that the PCD was an energy dependent active process and was controlled by nuclear genes. On the other hand, the distribution of ATPase in the intercellular spaces increased with the formation of the new cambium resulted from the dedifferentiation of the secondary xylem cells after girdling. However, ATPase was not found in the nucleus of the dividing cells, suggesting that nutrients were transported through protoplast during differentiation, and through both protoplast and apoplast during dedifferentiation. Thus, the energy required in cell division was provided mainly by intercellular spaces. These findings indicate that the dynamic distribution of ATPase reflected which cell component was actively taking part in the cell metabolism at various stages of the plant development, and its distribution was associated with the physiological state of the cell. Based on the characteristic distributions of ATPase, the critical stage of cell differentiation and the relationship between the critical stage and dedifferentiation were discussed.

Programmed Cell Death During Secondary Xylem Differentiation in Eucommia ulmoides

Programmed cell death (PCD) during secondary xylem differentiation in Eucommia ulmoides Oliv. was examined using electron microscopy and by investigation of DNA fragmentation and degradation of caspase-like proteases (CLPs). DNA ladders were detected in developing secondary xylem by gel electrophoresis. DNA fragmentation was further confirmed by using the TdT-mediated dUTP nick-end labeling (TUNEL) method. Western blotting analysis showed that CLPs (caspase-8- and caspase-3-like proteases) and PARP (poly (ADP-ribose) polymerase) were degraded during secondary xylem differentiation. The results thus indicated that secondary xylem differentiation in E ulmoides was a typical process of PCD and the degradation of CLPs might be a constitutive PCD event during secondary xylem differentiation.