Dynamic Changes Of Microtubules And Polysaccharide In Metaphloem Sieve Elements In The Developing Caryopsis Of Triticum Aestivum L.

Phloem sieve elements (SEs) in abdominal of wheat caryopsis play important roles in transport of photoassimilates.Previous studies suggested that the differentiation of metaphloem sieve elements (MSEs) in the developing caryopsis of wheat was an especial programmed cell semi-death process.And the concentration changes of Ca2+ and Ca2+-ATPase in MSEs might be related to the process.This study was carried on wheat (Triticum aestivum L. cv. Huamai 8).Transmission electron microscopy, indirect immunofluorescence labeling, periodic acid-Schiff/toluidine blue O double staining (PAS), periodic acid-Thiocarbohydrazide-silver proteinate method (PA-TCH-SP), and cellulase localization were used to observing phloem differentiation and MSEs development in the caryopsis of wheat. In addition, the dynamical changes of microtubules, polysaccharides and cellulase in MSEs were also studied. We discussed the relationship between those changes and the cell wall development, the cessation of PCD,and the photoassimilates transportation. The main results were as follows:1.Structural observation of phloem and MSEs showed that both the number of SEs and the area of phloem were rapidly increase from 0 to 8 days after flowering (DAF).The growth rate slowed down from 8 to 14 DAF. The nuclei of MSEs were normal at 0 DAF. From 2 to 4 DAF, the cell wall thickened unequally, and more conspicuous thickening appeared in corner of the cell; nuclei distortion and chromatin condensation were observed, and the mitochondria showed obscure cristae.Mature MSEs were short of cell contents, the cell wall became smooth, and a large number of cavities formed nearby the plasma membranes.The thickness of cell wall was small at 0 DAF. It increase quickly with MSEs developing and reached the peak at 4 DAF.After that, it slightly decreased and tended towards stability from 8 to 14 DAF.2.Immunofluorescence labeling of microtubule showed that the microtubules distrubuted in the middle side of two cells or around the cell wall at 0 DAF.The horizontal and network-like arrangement of microtubules appeared during MSEs ontognic development (from 2 to 3 DAF).At 4 DAF,the bipolar microtubules of cell increased. But the array of microtubules became vague with MSEs maturing at 6 DAF. The fluorescence of microtubules was not easy to be observed in the control. Sub-celluar observation showed that a few microtubules lay close to the plasmalemma at 0 DAF. Numerous microtubules distributed in the vicinity of the thickening cell wall or along the lateral wall at 2 DAF. From 3 to 4 DAF, a large of Golgi vesicles appeared; most of the microtubules closed to the cell wall of MSEs, they paralleled to each other and had the same direction as cellulose microfibril distribution. The microtubules and the vesicles disappeared gradually in matures MSEs.3.PAS staining of polysaccharide showed that the cytoplasm of few MSEs was dyed red by Schiff reagent from 1 to 4 DAF,and the cell wall had very slight coloration. At a later stage, the number of MSEs increased at 6 DAF;the cytoplasm of most MSEs was dyed red by Schiff reagent, and it maintained bright red till 15 DAF,but the cell wall could not be dyed red. The location of mucilaginous polysaccharides showed that there was no mucilaginous polysaccharide in MSEs from 3 to 35 DAF. But a spot of mucilaginous polysaccharides started to appear in the cytoplasm of chalazal at 7 DAF. From 10 to 25 DAF,more and more mucilaginous polysaccharides deposited in the vacuole of chalazal.At 35 DAF, mucilaginous polysaccharides did not increase any more. PA-TCH-SP reaction showed that the cell wall of MSEs always had a lot of sliver granules.There were still small glycoconjugates on the Golgi vesicles and nucleus during MSEs differentiation. A few states of aggregation silver granules deposited in the cytoplasm at 6 DAF.More and more state of aggregation silver granules appeared in the cytoplasm of mature MSEs till 15 DAF.4.Localization of cellulase showed that there was no cellulose on the plasmalemma of MSEs at 1 DAF.A few activity of cellulase distributed on the plasmodesmata during MSEs differentiation (from 3 to 4 DAF), and high levels of cellulase activity were observed on the plasmalemma of CC. At 6 DAF, a large number of cellulase activity appeared on the plasmalemma of matured MSEs.These results showed the dynamic changes of microtubules, polysaccharides, and cellulase were related to programmed cell semi-death process during MSEs differentiation in the developing caryopsis of wheat. They were in tight contact with the new cell wall formation, the cell wall non-uniform thickening, the cessation of PCD,and the assimilates transportation.