Final Activity Report Summary - RHIZO PHOSPHATASE (Optimising the efficacy of phosphatase in the rhizosphere, increasing the sustainability of agricultural crops)
At the outset of this project little was known about how effective phosphatase-organic P interactions in soils were with respect to plant phosphorus (P) nutrition. This project bought together a range of novel techniques including the use of transgenic plants, rhizosphere sampling and fluorescent labelled phosphatases to allow a completely new approach to this problem.
The research demonstrated that exudation of phosphatase by plants was related to mineralisation of specific forms of organic-P in soil and that plants which express genes for a specific phosphatase were able to deplete pools of phytate, the most abundant form of organic-P in soil. It was demonstrated that fertilisation of agricultural systems increased the content and availability of organic-P that was amenable to hydrolysis by phosphatase.
However, plants which express genes for a specific phosphatase were not able to acquire more P from such soils than control plants. One potential reason for this lack of effectiveness of specific phosphatases was a strong interaction between soil microorganisms and plant growth in P-limited conditions. When plants were grown in sterilised soil P-accumulation was improved, suggesting that soil microorganisms control P-turnover in soils. Moreover, expression of genes for a specific phosphatase was shown to be effective in sterilised soils but not in live soils. This suggests that where microorganisms exist there is no advantage to exuding specific phosphatase due to microbial production of phosphatases.
Another potential compromising factor was that phosphatases were rapidly adsorbed to the soil solid surfaces. Such adsorption was reversed by increasing pH, an effect which suggests that adsorption is dominated by electrostatic interactions and therefore the isoelectric point (pI) of the phosphatase protein was important. Subsequently, the research demonstrated that phosphatases with more acidic pI were more mobile in soil solution, more efficient at mineralising organic P, but were less stable in typical soil environments.
Phosphatase which remained in soil solution was shown to be more effective at releasing P from endogenous organic-P forms in a range of acidic soils and plants which expressed genes for this phosphatase grew larger in such soils. The culmination of this line of research was the observation that plants which expressed genes for phosphatases that remained in soil solution were particularly effective at acquiring P when grown on manured soils.
This implies that plants which exude phosphatases may be useful to deploy into systems which are reliant on fertilisation with animal manure. In other work up to 10-fold differences in exuded forms of various phosphatases were demonstrated in wheat and variability in this specific phosphatase signature was related to their ability to utilise organic forms of P under controlled conditions. However, this was a weak predictor of plant growth and nutrition in soils. Root associated phosphatase with activity against sugar phosphates was shown to be the best predictor, but this was extremely soil dependent and only accounted for a small proportion of the genotypic variability observed.
Another major outcome of this project was the production useful experimental material for future work into the spatial and temporal dynamics of phosphatase in soil. Constructs for gfp-phosphatase fusions have been produced and plants transformed. Plants expressing such genes have been verified to produce gfp fused with an active phosphatase, thus potentially allowing the visualisation of phosphatase in-situ in the rhizosphere.
Overall work completed in the course of this fellowship has demonstrated that root-associated phosphatase activity is a useful trait for improving plant P-nutrition but only under distinct environmental conditions where phosphatase remains in solution, where sufficient organic-P substrates exist and where the impact of microorganisms is weak.
The research demonstrated that exudation of phosphatase by plants was related to mineralisation of specific forms of organic-P in soil and that plants which express genes for a specific phosphatase were able to deplete pools of phytate, the most abundant form of organic-P in soil. It was demonstrated that fertilisation of agricultural systems increased the content and availability of organic-P that was amenable to hydrolysis by phosphatase.
However, plants which express genes for a specific phosphatase were not able to acquire more P from such soils than control plants. One potential reason for this lack of effectiveness of specific phosphatases was a strong interaction between soil microorganisms and plant growth in P-limited conditions. When plants were grown in sterilised soil P-accumulation was improved, suggesting that soil microorganisms control P-turnover in soils. Moreover, expression of genes for a specific phosphatase was shown to be effective in sterilised soils but not in live soils. This suggests that where microorganisms exist there is no advantage to exuding specific phosphatase due to microbial production of phosphatases.
Another potential compromising factor was that phosphatases were rapidly adsorbed to the soil solid surfaces. Such adsorption was reversed by increasing pH, an effect which suggests that adsorption is dominated by electrostatic interactions and therefore the isoelectric point (pI) of the phosphatase protein was important. Subsequently, the research demonstrated that phosphatases with more acidic pI were more mobile in soil solution, more efficient at mineralising organic P, but were less stable in typical soil environments.
Phosphatase which remained in soil solution was shown to be more effective at releasing P from endogenous organic-P forms in a range of acidic soils and plants which expressed genes for this phosphatase grew larger in such soils. The culmination of this line of research was the observation that plants which expressed genes for phosphatases that remained in soil solution were particularly effective at acquiring P when grown on manured soils.
This implies that plants which exude phosphatases may be useful to deploy into systems which are reliant on fertilisation with animal manure. In other work up to 10-fold differences in exuded forms of various phosphatases were demonstrated in wheat and variability in this specific phosphatase signature was related to their ability to utilise organic forms of P under controlled conditions. However, this was a weak predictor of plant growth and nutrition in soils. Root associated phosphatase with activity against sugar phosphates was shown to be the best predictor, but this was extremely soil dependent and only accounted for a small proportion of the genotypic variability observed.
Another major outcome of this project was the production useful experimental material for future work into the spatial and temporal dynamics of phosphatase in soil. Constructs for gfp-phosphatase fusions have been produced and plants transformed. Plants expressing such genes have been verified to produce gfp fused with an active phosphatase, thus potentially allowing the visualisation of phosphatase in-situ in the rhizosphere.
Overall work completed in the course of this fellowship has demonstrated that root-associated phosphatase activity is a useful trait for improving plant P-nutrition but only under distinct environmental conditions where phosphatase remains in solution, where sufficient organic-P substrates exist and where the impact of microorganisms is weak.