The Role Of Sulfate Assimilation Pathway In The Responses To Aluminum Stress In Arabidopsis

Aluminum toxicity has been recognized as a major abiotic factor limiting crop production in acid soils. Sulfur (S) is one of the essential macro-nutrient for plants, plays critical roles in response to biotic or abiotic stress. Thus, it is of great importance in studying the interaction between Al and S. However, little is known about the mechanisms of how Al affects S uptake and metabolism or the possible roles S plays in response to Al stress in plants.In the present study, we first investigated the expression patterns of key genes that function in sulfate (SO42-) uptake, translocate and reduction upon Al stress. After that, we carried a serious of physical experiments and reverse genetics study to analysis the interaction mechanisms between Al and S. The main results are as follows:1. Al3+ differentially regulates mineral nutrient acquisitionIn the present study here, we found that Al3+ inhibits absorption of K+, Ca2+ and Mg2+, among which the inhibition of Mg2+ uptake was the most severe; When it comes to Fe3+ acquisition, we found that Al3+ promote its accumulation in Arabidopsis roots in a big way; Al3+ significantly increased P accumulation in Arabidopsis roots, but reduced S content dramaticlly in Arabidopsis roots. These results indicate that the effects of Al3+ on the mineral nutrient acquisition were different among each other.2. Al3+ inhibits sulfate uptake by down-regulating the expression of SULTR1;1 and SULTR1;2In our study, the expression of SULTR1;1 and SULTR1;2 were quickly reduced upon Al3+ stress in WT Arabidopsis (Col-0), and it’s much more severe in Al3+ hypersensitive mutant, stopl. In our opinion, this should contribute to the inhibition of SO42- uptake. However, the expression levels of SULTR1;1 and SULTR1;2 were gradually restored after 3d under Al3+ stress. In addition, physiological results showed that SO42- content in roots after 7d treatment was not lower than 3d, and even with a slight increase, though total S content was still obviously lower than that at 3d. It is probably because the S level was at an adverse status after 3d treatment, the-S signal gradually surpassed over Al signal, thus induced the expression of SULTR1;1 and SULTR1;2 as well as the sulfate uptake in Arabidopsis roots.3. Al3+ activates the translocation of sulfate from roots to shoots as well as sulfate reduction pathwayWe found that the effect of Al3+ on total S or SO42- content in Arabidopsis shoots is different from that in roots. The former was at a relative stable level, while the later was reduced significantly. Moreover, it is also quite different form that under -S condition, which had a reduced S or SO42- level as time gone by. Thus, we proposed that Al3+ probably accelerate translocation of SO42- from roots to shoots. The expressions of transporter genes function in translocation of SO42- from roots to shoots were rapidly induced upon Al3+, which confirmed my speculation.As we know, most of the organic compounds in plants were produced mainly in shoots. In consideration of results above, we thought that it has something to do with the SO42- reduction pathway. Gene expression analysis showed that Al3+ quickly induced the expression of SULTR3;1, which facilitates transport of SO4- into chloroplast, as well as APR genes, key components in the SO42" reduction pathway. And finally it increased the biosynthesis of Cys and GSH, which plays a role in alleviation of Al toxicity, as results in in vitro experiment. Thus, we conclude that Arabidopsis has a mechanism to cope with Al toxicity and oxidative stress induced by Al through activation of SO42- translocation and reduction pathway and by improving the biosynthesis of thiols compounds like GSH.4. Synergetic acquisition of Al3+ and SO42- exists in response to Al stressAs it concluded above, Al3+ activated SO42- translocation and reduction pathway, while inhibited SO42- acquisition in Arabidopsis roots. It seems to be a conflict between SO42- supplies and needs upon Al3+ stress. It led me to think whether the inhibition of SO42- acquisition is also an adaptive mechanism under Al3+ stress.By testing Al3+ and SO42- accumulation in Arabidopsis roots in the SO42" does experiment (with 50μM AlC13 in the solution), we found the least Al and SO42-accumulation under -S condition. When the molar ratio of SO42- and Al greater than 1 (or SO42- concentration> 50μM), the content of Al in roots would not increase significantly, but with slight improvement, similar to SO42- uptake. Thus, there seems to be a synergetic mechanism exists in Al3+ and SO42- acquisition. In addition, the phenotype analysis and physiological experiment of overexpression lines of SULTR1;1 and SULTR1;2 confirmed this propose. Though increased SO42- acquisition in overexpression lines, the accumulation of Al were also improved upon Al3+ stress, exhibited hypersensitivity to Al3+. The inhibition of SO42- acquisition in Arabidopsis is therefore an adaptive mechanism when exposure to Al stress.5. Auxin play a role in Al accumulation and SO42- acquisitionAl3+ induced IAA biosynthesis in roots apex, and it’s the major factor that inhibits root elongation under Al3+ stress. PILS5 is a PIN-FORMED (PIN) auxin carrier regulates intracellular auxin homeostasis. Al quickly induced the expression of PILS5. The T-DNA insertion mutant pils5 showed significantly lower Al accumulation in roots compared to wild-type, while failed to exhibit a resistant phynotype. Higher free IAA levels in pils5 mutant compared to WT probably can explain this. In addition, the expression of SULTR1;1 and SULTR1;2 were significantly down-regulated in pils5 mutant compared to WT under both control or Al3+ stress condition, which must be the main reason why Al accumulation in pils5 mutant was deduced, as we known a synergetic acquisition of Al3+ and SO42- exists in response to Al3+ stress.In conclusion, the conflicts between SO42- supplies and needs indicate an adaptive mechanism in the early response to Al3+ stress in Arabidopsis. Cellular auxin homeostasis mediated by PILS5 play a role in the early response to Al3+ stress in Arabidopsis roots in the sulfate assimilation pathway.