The Mechanism Of Xanthophyll Cycle Regulation On Chlorophyll Metabolism Of Spinach Leaves Under Seawater Stress

China is short of freshwater resource, while is rich in beach and seawater resources. It seems that seawater cultivation can provide people with a a large of of the means of production and foodstuffs by taking advantage of its abundant beach and seawater resources. Seawater cultivation can inhibits plant growth as the formation of salt damage exacerbating photoinhibition, so it is very important to study the physiological and molecular biological mechanisms of crops under seawter stress, with significant theoretical and practical value and supply of general technique in taking advantage of saline-alkali soil. Firstly, the effects of seawater stress on chlorophyll metabolism was investigated using two cultivars plants cultured in the Hoagland’s nutrients solution with or without seawater (40%), the mechanism of xanthophyll cycle regulaiton on chlorophyll metabolism of Helan No.3(seawater tolerant cultivar) under seawater stress was investigated in the same method. Main research results were as follows:The contents of chlorophyll b(Chlb), chlorophyll a(Chla),Chl(a+b) and precursor of Chl such as protochlorophyll (Pchl), Mg-protoporphyrinⅨ(Mg-ProtoIX), protoporphyrinIX (Proto IX) and uroorphyrinogenⅢ (UroⅢ) in two spinach cultivars were decreased significantly, whereas Porphobilinogen (PBG) and8-Aminolevulinic acid (ALA) were increased significantly by seawater with higher rate in cv. Yuanye. The activity of chlorophyllase (Chlase) activity was not influenced in the leaves of Helan No.3under seawater stress, whereas activities of porphobilinogen deaminase (PBGD) and uroporphyrinogen Ⅲ synthase (UROS) were decreased significantly on the third day of treatment. As to cv.Yuanye, PBGD and UROS were decreased significantly with the Chlase activity increasing significantly. The results showed that chlorophyll biosynthesis in leaves of two spinach cultivars was blocked by seawater stress at the step from PBG to UroⅢ, and cv. Yuanye was more susceptible to seawater than Helan No.3. The decrease of Chl in cv. Helan No.3was caused by block of chlorophyll synthesis, whereas the reasons of the decrease of Chl in cv. Yuanye were block of chlorophyll synthesis and accelerating degradation of chlorophyll.Production rate of superoxide radical (O2-), content of hydrogen peroxide (H2O2) and malonaldehyde (MDA) were remarkably increased by seawater and methyl viologen (MV) which could induce oxidative stress on leaves of Helan No.3, whereas contents of chlorophyll a (Chla), chlorophyll b (Chlb), total chlorophyll and carotenoid (Car) remarkably decreased. Under the stress of seawater and treated with MV, maxmium quantum efficiency of photosystemⅡ photochemistry (Fv/Fm), the effective PSII quantum yield (Yield), apparent electron transport rate (ETR) and coefficient of photochemical quenching(qP) were decrease significantly, coefficient of nonphotochemical quenching(NPQ/4)was increased significantly. Accumulation of reactive oxygen species (ROS), degradation of photosynthetic pigments, as well as decrease of Fv/Fm, Yield, ETR, NPQ, qP were aggravated by introduction of dithiothreitol (DTT). Our results suggested that the inhibition of xanthophyll cycle under seawater stress decreased the dissipation of non-radiative energy, enhanced the accumulation of ROS, which were leading the degradation of photosynthetic pigments, decreation of activity of PSII and ETR, reduction of energy for photochemical reactions, and photosynthetic performances were affected significantly. Higher activity of xanthophyll cycle in seawater-tolerant spinach plays a major role in maintaining the stability of photosynthetic pigments and photosynthetic activity under seawater stress.On the third day of treatment, introduction of DTT in leaves of Helan No.3under seanwater stress further decreased the contents of Chlb, Chla, Chl(a+b) and precursor of Chl such as Pchl, Mg-ProtoⅨ, Proto IX and UroIII as well as the activity of PBGD with higher contents of PBG, ALAand activity of Chlase. The production rate of O2-, content of H2O2and MDA in chloroplasts of spinach were remarkably increased by DTT under seawater stress. Our results suggested that the inhibition of xanthophyll cycle under seawater stress decreased the dissipation of non-radiative energy, enhanced the accumulation of ROS, which were exacerbating the disorders of chlorophyll synthesis, promoting the further degradation of chlorophyll with the result of reduced chlorophyll contents. Higher activity of xanthophyll cycle in seawater-tolerant spinach plays a major role in maintaining the stability of chlorophyll metabolism under seawater stress. In the study, fragment of gene coding PBGD in spinach were cloned through RT-PCR, and its expressions under different treaments were analyzed using semi-quantitative RT-PCR. The results suggested the fragment of PBGD was389bp in length, encoding129amino acids which shared90%identities with it of Amaranthus tricolor. The expression of PBGD gene was inhibited by seawater stress which was aggravated by DTT. Our results suggested that the inhibition of xanthophyll cycle under seawater stress inhibited the expression of PBGD by the accumulation of ROS destorying the synthesis of chlorophyll.