| The vegetative storage proteins in subtropical fruit tree-Lychee (Litchi chinensis Sonn.) were identified by light- and electron microscope, SDS-PAGE, western-blotting and indirect immunohistochemical localization. On the basis ,we further investigated the distribution and seasonal changes of the vegetative storage proteins in Lychee by light microscopy and SDS-PAGE.In micrtome sections stained with mecury-bromophenol blue, abundant blue protein inclusions were present in many secondary phloem cells and xylem cells of the terminal branchlets. In the transverse section of branch, the protein inclusion-containing cells are the secondary phloem parenchyma cells in bark, and the various active parenchyma cells in xylem. So, we refered the cells as protein-storing cells. The protein-storing cells were also found in the secondary phloem cells of trunk and large roots, but poor in small roots and were not observed in the secondary xylem of trunk and large roots.Under the electron microscope, abundant electron dense materials in a floccular or lump appearance could be detected in the central vacuoles of the secondary phloem parenchyma cells, corresponding to protein inclusions observed under the light microscope. These vacuole proteins may accumulate firstly in small vacuoles, then in central vacuoles as the result of small vacuole combination.At the leaf-stable period, a relative abundance protein with molecular weight of about 22kDa was detected by SDS-PAGE, in the bark tissue of terminal branchlets,lateral branches and large roots. The protein also existed in large quantity in the secondary xylem of terminal branchlets and lateral branches, but was poor in the secondary xylem of trunk,large roots, and in small roots. The distribution pattern of 22kDa protein synchronized with that of vacuole protein inclusions inprotein-storing cells, suggesting that the 22kDa protein may be the main component of the vacuole protein inclusions.The 22kDa protein was purified by preparative SDS-PAGE and its polyclonal antibodies were obtained. Western-blotting analysis showed that the antibodies were highly specific to the 22kDa protein.We have further demonstrated the 22kDa protein to be the mam components of the vacuole protein inclusions of the protein-storing cells by indirect immunohistochemical localization. In the sections treated with the 22kDa protein antiserum, only the vacuole protein inclusions in the protein-storing cells gave off FTTC-specific green fluorescence.The seasonal fluctuation of the 22kDa proteins in bark tissue of autumn shoots was investigated during the development of new shoots, flowering and fruiting. The 22kDa protein accumulated in large amounts in the bark tissue of the first autumn shoots. When the second autumn shoots development, the 22kDa proteins stored in the first autumn shoots disappeared largely. The accumulation of 22kDa protein occurred in the bark tissues of first and second autumn shoots soon after the leaves of the second autumn shoots matured. At the early stage of floral bud differentiation, 22kDa protein existed in considerable amounts in both first and second autumn shoots. The protein decreased gradually in the late period of floral bud differentiation, blooming and fruit development, then no longer decrease at the late stage of fruit development. This seasonal pattern suggests that the 22kDa protein accumulating in branches play an important role in the growth and development of the new shoots, flowering and fruiting, and its hydrolysis products (amino acids) may be an important N resource for the development of new shoots, flowering and fruiting.In short, our results testify that a 22kDa protein is the vegetative storage proteinin Lytee tree. The protein mainly accumulated in branches, especially in the shoots which were the flowering and fruiting sites in future. The disapperance of 22kDa protein in shoots synchronized with new shoots growth, blooming and fruiting, suggesting that it was an important N resource to support the growth and development of new shoots,... |
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