| Acidic soil(pH<5.5)accounts for about 40%of the world's arable land,mostly in developing countries in Africa,Asia and South America.Aluminum is the main limiting factor for plant growth and crop yield in acid soils.The toxic effects of aluminum on plants are mainly manifested in the inhibition of root growth and development,and then affect the absorption of mineral nutrients and water.In the process of plant evolution,a variety of aluminum resistance mechanisms have been developed,and the organic acid anions secretion has been considered to be the most effective mechanism of aluminum resistance.Auxin is one of the most important hormones found in plants,which is involved in many physiological and stresses processes.The results of previous studies showed that exogenous IAA could significantly increase the lateral root development and citric acid secretion of soybean roots;however,the molecular and genetic mechanisms behinde the specific regulation are not clear.Using Arabidopsis thaliana as plant materials,the present study investigated the regulatory mechanisms of auxin channel protein PIN2 and AUX1 in organic acid anions secretion,root growth development and magnesium involved in the regulation of root development under aluminum stress.The results of the present study provided a theoretical basis for understanding the physiological and molecular mechanisms of plant response to A1 toxicity stress.The main results of this study are as follows:1.The present study firstly investigated the effect of aluminum on auxin accumulation and the regulation mechanism of PIN2 and AUX1 in organic acid secretion in Arabidopsis.The results showed that the fluorescence intensity of DR5::GFP was increased with the increase of aluminum concentrations,indicating that Al induced IAA accumulaton in an Al-dependent manner.Studies on fluorescence confocal results showed that Al did not induced the expression of PIN3;however,the fluorescence intensities of PIN2 and AUXI were obviously increased.Coincidently,RT-PCR analysis showed that the expression of PIN2 and AUX1 was increased by 5.11 and 4.69 times comparing with the control treatments.These results indicated that A1 induced IAA accumulation in Arabidopsis root may due to upregulation of PIN2 and AUX1.We then analyzed the regulation mechanism of PIN2 and AUX1 on the secretion of organic acids under aluminum stress.The results showed that the secretion amonnts of citrate in the roots of pin2 and aux1-7 mutants were significantly decrased while comparing with the Col-0 plants under A1 stress.However,there were no significant changes in malic acid contents.Compared with the control treatment,the apoplastic pH values in the pin2 and aux1-7 in were increased by 12.26%and 10.53%under A1 stress.These results indicated that PIN2 and AUX1 are involved in Al-induced citrate exudation through regulating the activity of proton pump.2.We further analyzed the regulation mechanisms of PIN2 and AUX1 on root development of Arabidopsis under aluminum stress.The study found that the 50 ?M Al treatment of Arabidopsis after 5 days,compared with the control treatment,the root elongation was inhibited by 47.70%;However,the number of lateral roots increased by 23.86%,A and B period were reduced by 30.12%,61.90%,C period increased by 5%,the number of D period increased 4.36 times;the number of lateral roots in the period of A,B period was reduced by 56.25%,31.03%,in the period of C,D were increased by 96.67%,41.67%in the proximal;the number of lateral roots in the period of A,B was reduced by 62%,57.14%,in the period of C,D were increased by 87.5%,60.42%in the distal,and GUS staining analysis also found that the lateral root auxin content increased.The above results indicate that aluminum can inhibit the growth of primary root but promote lateral root elongation by inducing the accumulation of IAA.Research on the pin2 and auxl-7 mutants of lateral root development of the results showed that compared with Col-0,pin2 and auxl-7 mutants of root elongation increased by 33.33%and 18.52%,but the number of lateral roots was reduced by 18.18%and 34.09%under aluminum stress.Among them,the number of proximal lateral roots increased by 11.59%and 18.84%,and the number of distal lateral roots decreased by 26.67%and 60%.Aluminum treatment,compared with Col-0,the number of root hairs pin2 and auxl mutant were reduced by 29.13%,48.92%,root hair length was reduced by 41.17%54.24%,indicating that PIN2 and AUX1 can regulate the development of root hairs,by promoting the development of root hairs in response to aluminum stress.Thus,PIN2 and AUX1 mediated IAA in root involved in the accumulation of aluminum stress on arabidopsis root growth inhibition and lateral root and root hair growth promotion.3.Magnesium is an important nutrient element in plants,which not only plays a key role in plant growth and development,but also participates in the response of plants to Al toxicity.However,the mechanism of the regulation of magnesium on the development of plant roots under aluminum stress is not clear.Therefore,we investigated the role of PIN2 and AUX1 in the regulation of root development in Arabidopsis plants under A1 stress.The results show that,compared with not adding magnesium,adding 10,50,300,500,1000?M of magnesium DR5::GUS root elongation were increased by 33.69%,39.52%,35.46%,37.11%,2.33%;however,2000?M magnesium treatment,root elongation was inhibited by 55.55%.Study found that,compared with the control group(CK,Al0Mg0),adding 50?M of aluminum,the Arabidopsis root elongation inhibition by 45.01%,however,compared with the treatment without adding magnesium(MgoAlso),adding 10,50,300,500 and 1000?M of magnesium root elongation were increase by 59.05%,55.21%,40.79%,31.83%,11.05%,and after 2000?M magnesium treatment root elongation was inhibited by 73.75%.The analysis results of lateral roots showed that adding 10?M Mg after the number of lateral roots increased by 14.12%,while adding 50,100,300,1000,2000?M Mg the number of lateral roots was decreased by 2.94%,6.86%,17.65%,52.94%,83.33%,indicating that low concentration of magnesium could alleviate the inhibition of Al on the growth of primary root and promote the growth of lateral roots,high concentration inhibited growth.Subsequently,we investigated the effect of exogenous addition of magnesium on cell division in quiescent central and meristematic regions under aluminum stress.The results showed that after treatment with aluminum,QC::GUS plants and CycB1,1::GUS the color of the plants was lighter after GUS treatment;However,after the addition of 10?M of magnesium,the inhibition of on the expression of QC and CycB1 was reversed by the addition of aluminum.The results indicated that magnesium could effectively alleviate the inhibition of A1 toxicity on the development of stem cells and the division of meristematic cells.Finally,we analyzed the effects of PIN2 and AUX1 on root growth of Arabidopsis thaliana under A1 stress.The results showed that the addition of 10?M of magnesium promoted the expression of AUX1,but the expression level of PIN2 decreased.The analysis of the primary root growth showed that exogenous addition of magnesium under Aluminum Stress,compared with Col-0 the root elongation of pin2 mutant was increased by 17.67%,but the main root of aux1-7 was not affected by magnesium treatment.The staining results of DR5:GUS/pin2 and DR5:GUS/aux1 showed that the addition of magnesium inhibited the accumulation of auxin in the root tip of pin2 mutant,while the content of auxin in AUX1 mutant was increased.In conclusion,adding Mg can alleviate the toxicity of A1 to plants,mainly due to exogenous magnesium can effective regulate PIN2 and AUX1 expression level to regulate the balance of IAA in Arabidopsis root tip under aluminium stress.Finally,the effect of aluminum on the development of root tip stem cells and the division of meristematic cells in meristematic zone was inhibited,the growth of main root and lateral root development were maintained. |
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