Effects On The Tubulin And Microtubule Cytoskeleton Of Wheat Seedlings Exposed To Enhanced Ultraviolet-b Radiation And Irradiated He-Ne Laser

Taken the wheat seedlings of Jin mai No.8 as material, which were exposed to He-Ne laser irradiation (5mW·mm-2), enhanced UV-B radiation(10.08kJ·m-2·d-1) and the combined treatments of its both. In order to study the effects of UV-B and He-Ne laser on the tubulin and microtubule, as well as to further investigate relationships between microtubule and'Partition-bundle division',the changes of tubulin content and microtubule arrangement were researched through indirect immunofluorescence localization and confocal laser scanning microscopy(CLSM). The results were showed as follows:1. Extract the tubulin by acetone from wheat leaves and roots, analysed protein of SDS-PAGE. The main brand was a kind of protein whose molecular weight is 50 kD.2. The protein content of tubulin was decreased under the condition of enhanced UV-B radiation. However, the tubulin content was increased in the treatments of UV-B radiation and He-Ne laser. This result showed that He-Ne laser stimulate the tubulin synthetic, and can repair the reduction of tubulin.3. The changes about wheat leaves'tubulin contents at different days were mesured, it showed that 8-day-old was the most content and has significant difference with others. To facilitate the looking for the best days of microtubule immunofluorescence localization in Mesophyll protoplasts.4. Mesophyll protoplasts in control group was spherical and smooth surface, but the UV-B group was stick , several cells were linked too. This demonstrated that the cells surface was changed, cell wall thicken, wax of surface increased as well. As a result of , enhanced UV-B radiation influenced the enzymatic effects.5. Study the microtubule in mesophyll protoplasts by immunofluorescence localization, results showed that microtubule of protoplast in wheat seedlings was depolymerized significantly to stick and spot under the condition of enhanced UV-B radiation, the MT bundles were trend to dispersivity and decreasing of fluorescence intensity . However, the depolymerization of MT is decreased by He-Ne laser irradiation after enhanced UV-B radiation. Therefore, the damage of microtubule in wheat seedlings induced by enhanced UV-B radiation can be repaired partly by He-Ne laser irradiation.6. The microtubules cytoskeleton of wheat root tips were labled with indirect immunofluorescence localization, the results of our study show that the microtubule arrays presented a periodicity during mitosis, as well as the obvious microtubule cycle and high fluorescence intensity were observed in control group . There were different microtubule arrangements in cell division cycle what namely microtubule cycle , including cortical microtubule of interphase, preprophase band (PPB) and the preliminary formation of spindle microtubules in prophase, spindle microtubules and phragmoplast microtubules in metaphase and anaphase, and phragmoplast microtubules in telophase cytokinesis, and its transitional arrangements.7. Under the condition of enhanced UV-B radiation, the structure of microtubules were disorder while MTs appeared a lot of abnormal arrangements. The cortical microtubules changed orientation to network and depolymerized significantly to stick and spot ; the cells had abnormal structure of double preprophase bands; the arrangement of spindle microtubules showed accumulation and the phragmoplast microtubules were dispersivity or disappeared due to enhanced UV-B radiation.8. The damage structure of spindle or gathered into a stick because of UV-B radiation will directly impact on the combination of chromosomes and spindles, while the changed structure of MTs will influence chromosome distribution dramatically. This so easy to cause chromosome accumulation or loses in cells, and lead to lagging chromosome, chromosome bridge, evev the'Partition-bundle division'. Additionally, the cells in telophase without clearly phragmoplast due to enhanced UV-B radiation, thus beyond control the cell plate material deposition and daughter cells can not constitute a new cell wall, lead to the phenomenon of multi-nuclear cells eventually.Microtubules are an essential menber of cell's cytoskeleton, it has become a hot research in field of life sciences since it was discovered. Information on the microtubule in higher plants has increased rapidly during the last couple of years, however, there have no reports about the influence of microtubule exposed to ultraviolet-B radiation and He-Ne laser. Our study taking advantage of indirect immunofluorescence localization and confocal laser scanning microscopy to analyse of combining tubulin and microtubules arrays, in order to investigate the effects of UV-B and He-Ne laser on microtubules dynamic in wheat cells, as well as the microtubule function in the plant defense mechanisms to enhanced UV-B. Under the condition of UV-B treatment, wheat root-tips were induced to appear abnormal mitosis that was'Partition-bundle division',in its cells, chromosomes were divided into different bundles, as viewed from the number of chromosomes in each bundle , it was smaller than that in normal somatic cells. Taking root-tips as material, the study aim is to observe the effects of UV-B radiation on the microtubule cycle and further to investigate the relationships of microtubules arrangement and'Partition-bundle division', while establish a basis for studying the mechanism of abnormal mitosis, the biological effects of UV-B as well.To sum up, the studies which effects on microtubule by UV-B radiation and He-Ne laser from cell biology have great significance, it provide theoretical basis for agriculture, forestry production and research of MTs at the molecular.