Morphological changes and the timing of disappearance of individual organelles provide key information for understanding the mechanism of cell death. The disappearance of microtubules, nuclei and starch grains was monitored during the death of long-lived ray parenchyma cells in the conifer Abies sachalinensis. From the eighth to the tenth annual ring from the cambium, morphological changes occurred in ray parenchyma cells and organelles disappeared. Morphological changes in nuclei became apparent first. Then microtubules disappeared and, finally, nuclei disappeared. Therefore, microtubules might play an important role during the death of ray parenchyma cells. The timing of the disappearance of starch grains differed among individual ray parenchyma cells. This result indicates that the timing of the loss of storage function in ray parenchyma cells might not depend on the progress of cell death. The possible role of microtubules during cell death of long-lived ray parenchyma cells that might differ from cell death of short-lived ray tracheids is discussed.
The aim of the present study was to investigate the orientation and localization of actin filaments and cortical microtubules in wood-forming tissues in conifers to understand wood formation. Small blocks were collected from the main stems of Abies firma, Pinus densiflora, and Taxus cuspidata during active seasons of the cambium. Bundles of actin filaments were oriented axially or longitudinally relative to the cell axis in fusiform and ray cambial cells. In differentiating tracheids, actin filaments were oriented longitudinally relative to the cell axis during primary and secondary wall formation. In contrast, the orientation of well-ordered cortical microtubules in tracheids changed from transverse to longitudinal during secondary wall formation. There was no clear relationship between the orientation of actin filaments and cortical microtubules in cambial cells and cambial derivatives. Aggregates of actin filaments and a circular band of cortical microtubules were localized around bordered pits and cross-field pits in differentiating tracheids. In addition, rope-like bands of actin filaments were observed during the formation of helical thickenings at the final stage of formation of secondary walls in tracheids. Actin filaments might not play a major role in changes in the orientation of cortical microtubules in wood-forming tissues. However, since actin filaments were co-localized with cortical microtubules during the formation of bordered pits, cross-field pits and helical thickenings at the final stage of formation of the secondary wall in tracheids, it seems plausible that actin filaments might be closely related to the localization of cortical microtubules during the development of these modifications of wood structure.
To understand the precise process of wood formation, it is necessary to identify the factors that regulate cambial activity and development of cambial derivatives. Here, we investigated the combined effects of localized-heating and auxin on cambial reactivation and the formation of earlywood tracheids in seedlings of the evergreen conifer Abies homolepis in winter. Three treatments were applied, namely heating (artificial increase in temperature 20–22 °C), heating-plus-auxin transport inhibitor N-(1-naphthyl) phthalamic acid (NPA) and heating-plus-defoliation (removal of needles and buds), with an approximate control, for investigations of cambial activity by light microscopy. After one week of heating, cambial reactivation occurred in the heating, heating-plus-NPA and heating-plus-defoliation treatments. In untreated controls, cambial reactivation occurred later than in heated stems. Earlywood tracheids were formed after three and six weeks of heating in the heating and heating-plus-NPA treatments, respectively. No tracheids were formed after eight weeks of heating in heated-defoliated seedlings. Numbers of new tracheids were reduced in heated stems by NPA. Our results suggest that an increase in the temperature of the stem is one of the most important limiting factors in cambial reactivation, which is independent of needles and buds and of the polar transport of auxin from apical sources. However, after cambial reactivation, initiation and continuous formation of earlywood tracheids require basipetally transported auxin and other endogenous factors originating in mature needles and buds.