Fine Root Senescence Of Manchurian Ash Seedlings Under Drought Stress

Fine roots are the most important components of tree root system,and have significant physiological and ecological functions.The development and tumover of fine roots have vital effects on the development of an individual tree,the forest productivity and the flux of material and energy in a forest ecosystem.Fine roots senescence is the transition phase of fine roots turnover.Its occurrence and processes are the key points of individual root development,root longevity and root turnover.Drought stress is one of the environment factors inducing fine root senescence.In order to make clear the real physiological responses of fine roots to drought stress and to explore the mechanisms of fine root senescence,one year old seedlings of Fraxinus mandshurica were used to investigated the outer morphological changes(including the color and diameter of root),the physiological changes(including root vigor,membrane permeability,soluble protein content and activities of antioxident enzymes),the inner structure changes(including microstructrue and ultrastructure),and DNA degradation.The aims of the present dissertation are to make more perfect the theory of fine root biology and to provide more base for fine root ecological research.The main results are summarized as follows:(1)There were five root orders in the root system of one years old seedlings of Fraxinus mandshurica,different order roots with different anatomical structure characteristics.All forthand fifth-order roots and a majority of third-order roots with successive phellems were second growth roots without ability to absorb water and nutrition,but with transport ability.All the first- and second-order roots and a small quantity of third-order roots,which diameter were less than 0.4 mm and without successive phellems,were the primary roots with absorption ability. These roots were defined as fine roots of one years old Fraxinus mandshurica seedlings.(2)Four colors of fine roots under drought stress,i.e.,white,yellow,brown and white root with black tips,were observed.They were more complex than the control that had just white and yellow roots.During the drought stress treatment,the numbers of white and yellow roots decreased but the brown and white roots with black tips increased gradually.The first-order roots under drought stress showed 4 colors mentioned above and other transitional colors.The second-oder roots showed 3 colors,i.e.,white,yellow and brown,which had the same trendancy as the first-order roots,there were yellow and brown roots in The third-order roots showed yellow and brown,but didn't show white.Therefore,the first-order roots were the most sensitive to drougth stress.(3) The average diameters of the first three order roots were significantly smaller than control(P＜0.05),and decreased during drought stress treatment.The decrease in the average diameters of the first-order roots was more than the second- and third-order roots.During the drought stress treatment,the diameters of different color roots changed differently.Compared with the control,the diameters of white first-order roots decreased most significantly.After 30 days of drought stress,the diameters of the second-order roots with three colors were significantly lower than control(P＜0.05).The diameters of yellow third-order roots changed little during drought stress.Dcreased diameter was the one of the typical morphological responses to drought stress for fine roots of Fraxinus mandshurica seedling.(4)During drought stress treatment,the root vigor and soluble protein content decreased, and the membrane permeability increased.There were significant relationships among these three physiological indexes(γ²=0.61-0.87,P＜0.05).The activities of the antioxidant enzymes in the first-order roots were higher than control,but their activities in the second-order roots were not.And the decline extents of the enzymatic activities in the first-order roots were higher than those in second-order roots druing drought stress treatment.It was obvious that the the possibility that membranes were oxidated occurred easier in first-order roots than in the second-order roots under drought stress,thus the first-order roots senesced at first under drought stress.(5) All the first- and second-order roots,and parts of third-order roots browned during drought stress treatment.The first- and second-order browned roots were primary roots with diarch protoxylems but without phellem.But the browned third-order roots were second growth roots with successived phellem and tetrarch protoxylems.The reason why the first- and second-order roots browned was that many cortical parenchyma cells died.And these were senescent roots induced by drought stress treatment.The quantity of dead cortical parenchyma cells in the first-order roots were higher than that in the second-order roots.Accorder to the anatomical analyses,there were two reasons for browning of the third-order roots.One reason was that a few successived phellem and more dead corical parenchyma cells occurred in the third-order roots.The parenchyma cells in phloem were still alive in the roots,and they still had ability to transport water and nutrition.The other reason was that there were so many successived phellem and lots of dead cortical parenchyma cell in the browned roots.These roots had neither transprot ability nor absorption ability,and they were senesceing under drougth stress.Therefore,it can be concluded that all the first-and second-order roots with diarch protoxylems and parts of third-order roots with dead cortical parenchyma cell and dead phloems senesced after drought stress treatment.(6)The shapes,structures and quantity of mitochondria in cortical parenchyma cells of white and yellow roots under drought stress were same as those of control.For few yellow roots under drought stress,there were lower center electron density in mitochondrias and fewer cristaes in their inner membrane.Compared with white and yellow roots,the browned roots induced by drought stress had fewer mitochondrias,which distributed seperately.The inner membranes of some mitochondrias with intact outer membrane had disorganized to some extent.But outer membranes of some mitochondrias with intact inner membrane also degraded. At last,the inner and outer membranes in the mitochondrias all dissapered and all the mitochondias decomposed.The respiratory control ratio of mitochondiras was strongly influenced by drought stress.The values of RCR and P/O in fine roots under drought stress were significantly lower than those in control,and decreased significantly with root colors,i.e., white root＞yellow root＞brown root.The RCR value of the brown roots was the lowest, close to 1.All these results indicated that the integrality of mitochondia in brown roots was the worst and they had little ability to produce energy.When mitochondria integrality was destroied,which resulted in the loss of respiration ability,fine roots senesced.(7)The nucleuses in the cells of white and yellow roots under drought stress were the same as those of control,with intact outline and a big nucleolus.The chromatin distribution of in the white and yellow roots of control assembled apparently and uniformly.It was few chromatin assembling in the white and yellow roots under drought stress.There were great changes in nucleus shapes in brown roots and black tips of white roots and those nuclear membranes gradually degraded and the nucleoluses disappeared gradually.The chromatin concentrated deeply but did't become chromosomes.As nuclear membranes and nucleoluses disappeared,the chromatin disappeared in the cytoplasm.The nucleus changes happening in brown and black tips of white roots were same as those happening in leave and animal cells' programmed cell death.On the other hand the DNA ladder and TUNEL test provided evidence that there were PCD happening in brown and black tips of white roots.There were no DNA ladder and TUNEL fluorescence signs in control roots.Althought there were not apparent DNA ladder and TUNEL fluorescence signs in yellow and white roots of control,there still a little TUNEL fluorescence signs seen in some chromatin,suggesting that some DNA molecule chain degraded.It was the premonitory behavior for PCD.In conclusion,drought stress induced the respiration reactions of mitochondria,which producing more H₂O₂.Some H₂O₂ couldn't be scavenged in time and induced oxidation of the mitochondria membranes so that mitochondria structures were detroied and mitochondria disappeared.As mitochondria structures and functions changed,the nucleus and DNA structure also changed.And then the PCD happened.The dead cells with more phellem assembled in the cell walls resulted in browning of fine roots.And at the same time the root vigor and content of soluble proteoins decreased,and membrane pemeability increased.At last the fine roots senesced.The lower order roots was more affected by drought stress than the higher order roots and senesced earlier.