Responses Of Maize Seedlings To Waterlogging And Its Regulation Mechanisms By Exogenous Plant Growth Regulators

Waterlogging is one of major abiotic stress on maize in sub-tropical and tropical region,which has serious threat to growth and productivity of maize.Vegetative stage of maize seedling development is considered extremely sensitive to waterlogging stress.However,susceptible growth stages of maize seedlings under waterlogging are not still fully clear,and the countermeasures to enhance maize seedlings endurance to waterlogging are in urgent need.Here,we conducted a series of experiments to knockout the morphological,physiological,anatomical,biochemical mechanisms at different growth stages of maize seedlings under waterlogging,and identify the modulatory effect by exogenous plant growth regulators application,then to reveal the physiological and biochemical potentials in fortifying the tolerance of maize seedlings to waterlogging.The main findings of these works are summarized below.1.The susceptible growth stages of maize seedlings were distinguished by using three maize cultivars,viz.HZ-32,Xingken-6?XK-6?and Zhengdan958?ZD-958? under waterlogging conditions?WL?initiated at V2,V3,V4,V5 and V6 stages in pot experiment,by contrasting the normal soil water treatment?CK?.The results showed that more reduction in the photosynthetic net rate,shoot dry matter and leaf area of maize seedlings under waterlogging?P<0.05?occurred at V2,V3 and V4 stages than V5 and V6 stages averaged across three cultivars.Similarly,great decrease in root dry matter under waterlogging was observed at V2 and V3 stages in comparison with other growth stages?P<0.05?.However,the highest drop in root length,surface area and volume of maize seedling under waterlogging were uniformly occurred at V2 stage.Regarding the aboveground growth,the cultivar of HZ-32 exhibited a relatively higher tolerance to waterlogging.2.This study investigated the effects of exogenously applied spermidine?SP?,brassinosteroid?BR?and aminobutyric acid?GABA?with determined optimal concentration on maize seedlings under waterlogging conditions.Two maize cultivars viz.Xingken-6?XK-6?and Zhengdan-958?ZD-958?were used under waterlogging treatments initiated at third-leaf?V3?and fifth-leaf stages?V5?in 2016 and 2017 repeated pot experiments.Chloroplast ultrastructure,photosynthesis,antioxidant capacity and reactive oxygen species?ROS?production in leaves under waterlogging with the plant growth regulators?PGR?were examined in compare with the normal soil water treatment?CK?and waterlogging treatment?WL?.The results revealed that PGR treatments could enhance the seedling growth under 14 days waterlogging stress.Compared with WL treatment,application of SP,BR and GABA showed constantly significant increase in shoot dry matter by 25%,44% and 34%,promoted total chlorophyll content by 20%,46%and 34%?P<0.05?,respectively.Moreover,the application of PGR also enhances the photosynthesis performance and effectively palliated the damage of ultrastructure of chloroplast.Furthermore,PGR application significantly reduced the level of ROS,which are related to the activities of ROS producing enzymes and noticeably up-regulated the activities of antioxidant and non-antioxidant enzymes in leaves under waterlogging stress.Among PGR treatments,the BR and GABA was the most effective treatments for both maize cultivars under waterlogging stress.3.On the basis of selection on PGR,root growth,anatomical structure and its related biochemical changes under BR and GABA application were emphatically examined in the two-year experiments.The results demonstrated that waterlogging stress significantly triggered ROS-production in roots,and remarkably increased aerenchyma area and formation,destroyed root structure,and eventually inhibited the morphological growth traits of maize seedling.However,exogenous application of BR and GABA enhanced adaption of maize root to waterlogging stress by reducing ROS-production,aerenchyma formation and promoting number of living cortical cell and antioxidant capacity.Remarkably,compared with WL treatment,on average of values across two varieties,stages and years,BR and GABA treatments constantly increased root dry matter by 81% and 57%,root length by 86% and 61%,root volume by 58%and 65%;number of living cortical cell by 46%and 31%?P<0.05?,respectively.Meanwhile,BR and GABA treatments significantly reduced the aerenchyma formation by 52%and 35%,suppressed ROS-production ?H₂O₂ and^·O₂^-?content by 69.25%and 46.15%?P<0.05?,respectively.BR application showed better effect on root growth of V3 maize seedlings under waterlogging than GABA.4.In order to improve our understanding of the complex mechanism of BR application underlying waterlogging stress and identify waterlogging responsive proteins,the iTRAQ LC-MS/MS was conducted to map the proteomes of maize seedlings of ZD-958 under BR treatment,WL and CK.A total of 4235 proteins were identified by iTRAQ,and there were 257,110 and 519 proteins were up-regulated in BR vs CK,WL vs CK and BR vs WL respectively.Among all,509,760 and 592 were identified as differential abundance proteins?DAPs?in BR vs CK,WL vs CK and BR vs WL respectively.Further,Gene ontology?GO?analysis suggested that different treatments demonstrated dynamic molecular mechanisms in their waterlogging responses.Eleven proteins were identified which were highly up-regulated and involved to redox homeostasis.Our result suggests that exogenous application of BR under waterlogging stress may rescue the plant and activated the plant cell defense by regulating defense responsive proteins.Conclusively,waterlogging had considerable adverse effects on both aboveground and root growth of maize seedlings,especially at more susceptible V2 and V3stages.However,PGR application,such as BR and GABA could obviously promote growth and root structure integrity of maize seedlings under waterlogging by activation of antioxidant defense systems,improving chloroplast ultrastructure and photosynthetic traits and reducing the aerenchyma formation.Moreover,exogenous BR and GABA application could protect maize seedlings from oxidative stress caused by waterlogging stress,due to inhibition of ROS-production through up-regulating proteins involved in redox homeostasis,which might be related to controlling the aerenchyma formation in seminal root.However,further investigation is required to better understanding regulative mechanisms by PGR on maize seedlings under waterlogging stress.