AM plants acquire Pi via two pathways; the direct uptake pathway via plant roots and the AM pathway via external fungal hyphae and colonised cortical cells. It has been assumed that these two pathways are additive and therefore in non-responsive plants the AM pathway is often considered to be non-functional. However, data from ³²P uptake studies indicates that the AM pathway is functional in many non-responsive symbioses and in some instances supplies the majority of plant P. In recent years the high-affinity Pi transporters involved in both direct and AM Pi uptake pathways have been identified. They are expressed at the root epidermis and the symbiotic interface of colonised cortical cells and respond to the P and AM status of the plant. The overall objective of the work described in this thesis was to characterise Pi uptake via the AM pathway in barley, a non-responsive AM host, using an approach which integrated physiological measurements of plant responsiveness and AM contribution with investigations of gene expression and functional characterisation of the plant Pi transporters. A preliminary survey of field-grown barley demonstrated the persistence of AM colonisation under commercial cropping regimes in southern Australia and highlighted the relevance of AM studies to commercial agriculture. Under glasshouse conditions AM colonisation of barley induced depressions in growth and P uptake compared to NM controls. Growth depressions were unrelated to percent colonisation by two AM fungal species and could not readily be explained by fungal C demand; the strong correlation between growth and P content suggested that P was the limiting factor in these experiments. However, a compartmented pot system incorporating ³²P-labelling demonstrated that the AM pathway is functional in colonised barley and, in the interaction with G. intraradices, contributed 48% of total P. This suggested that P flux via the direct uptake pathway is decreased in AM barley. The expression of three Pi transporters, HvPT1, HvPT2 and HvPT8 was investigated in colonised roots. HvPT1 and HvPT2 have previously been localised to the root epidermis and root hairs and are involved in Pi uptake via the direct pathway whilst HvPT8 is an AM-inducible Pi transporter which was localised by in-situ hybridisation to colonised cortical cells. Using promoter::GFP gene fusions the localisation of HvPT8 to arbuscule-containing cortical cells was confirmed in living roots from transgenic barley. Quantitative real-time PCR analysis of the expression of these three Pi transporters indicated that HvPT1 and HvPT2 were expressed constantly, under all conditions regardless of AM colonisation status and indicated that decreased P flux via the direct pathway is not related to expression of these transporters. HvPT8 was induced in AM colonised roots. However, the level of expression was not related to flux via the AM pathway or arbuscular colonisation. The HvPT8 transporter was further characterised by constitutive over-expression in transgenic barley. ³²P uptake assays in excised roots demonstrated increased Pi uptake from low P solution compared to wild-type roots and confirmed that HvPT8 is a functional Pi transporter with high-affinity transport properties. This is the first report of characterisation of an AM-inducible Pi transporter in planta. When these transgenic plants were grown in solution culture there was no increase in growth or P uptake relative to wild-type or transgenic controls and growth in soil and AM colonisation were also unaffected in these transgenic lines. The data presented in this thesis highlights the importance of combined physiological and molecular approaches to characterising plant AM interactions. The persistence of AM colonisation in barley in the field indicates the importance of improving our understanding of symbiotic function in non-responsive plants. Future efforts should be directed towards understanding the signals which regulate P flux via both the direct and AM pathways with the ultimate aim of enhancing AM responsiveness of non-responsive species. Making the direct and AM pathways additive in nonresponsive species should be a key aim of future research. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1313311 / Thesis (Ph.D.) -- University of Adelaide, School of Earth and Environmental Sciences, 2008

AM plants acquire Pi via two pathways; the direct uptake pathway via plant roots and the AM pathway via external fungal hyphae and colonised cortical cells. It has been assumed that these two pathways are additive and therefore in non-responsive plants the AM pathway is often considered to be non-functional. However, data from ³²P uptake studies indicates that the AM pathway is functional in many non-responsive symbioses and in some instances supplies the majority of plant P. In recent years the high-affinity Pi transporters involved in both direct and AM Pi uptake pathways have been identified. They are expressed at the root epidermis and the symbiotic interface of colonised cortical cells and respond to the P and AM status of the plant. The overall objective of the work described in this thesis was to characterise Pi uptake via the AM pathway in barley, a non-responsive AM host, using an approach which integrated physiological measurements of plant responsiveness and AM contribution with investigations of gene expression and functional characterisation of the plant Pi transporters. A preliminary survey of field-grown barley demonstrated the persistence of AM colonisation under commercial cropping regimes in southern Australia and highlighted the relevance of AM studies to commercial agriculture. Under glasshouse conditions AM colonisation of barley induced depressions in growth and P uptake compared to NM controls. Growth depressions were unrelated to percent colonisation by two AM fungal species and could not readily be explained by fungal C demand; the strong correlation between growth and P content suggested that P was the limiting factor in these experiments. However, a compartmented pot system incorporating ³²P-labelling demonstrated that the AM pathway is functional in colonised barley and, in the interaction with G. intraradices, contributed 48% of total P. This suggested that P flux via the direct uptake pathway is decreased in AM barley. The expression of three Pi transporters, HvPT1, HvPT2 and HvPT8 was investigated in colonised roots. HvPT1 and HvPT2 have previously been localised to the root epidermis and root hairs and are involved in Pi uptake via the direct pathway whilst HvPT8 is an AM-inducible Pi transporter which was localised by in-situ hybridisation to colonised cortical cells. Using promoter::GFP gene fusions the localisation of HvPT8 to arbuscule-containing cortical cells was confirmed in living roots from transgenic barley. Quantitative real-time PCR analysis of the expression of these three Pi transporters indicated that HvPT1 and HvPT2 were expressed constantly, under all conditions regardless of AM colonisation status and indicated that decreased P flux via the direct pathway is not related to expression of these transporters. HvPT8 was induced in AM colonised roots. However, the level of expression was not related to flux via the AM pathway or arbuscular colonisation. The HvPT8 transporter was further characterised by constitutive over-expression in transgenic barley. ³²P uptake assays in excised roots demonstrated increased Pi uptake from low P solution compared to wild-type roots and confirmed that HvPT8 is a functional Pi transporter with high-affinity transport properties. This is the first report of characterisation of an AM-inducible Pi transporter in planta. When these transgenic plants were grown in solution culture there was no increase in growth or P uptake relative to wild-type or transgenic controls and growth in soil and AM colonisation were also unaffected in these transgenic lines. The data presented in this thesis highlights the importance of combined physiological and molecular approaches to characterising plant AM interactions. The persistence of AM colonisation in barley in the field indicates the importance of improving our understanding of symbiotic function in non-responsive plants. Future efforts should be directed towards understanding the signals which regulate P flux via both the direct and AM pathways with the ultimate aim of enhancing AM responsiveness of non-responsive species. Making the direct and AM pathways additive in nonresponsive species should be a key aim of future research. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1313311 / Thesis (Ph.D.) -- University of Adelaide, School of Earth and Environmental Sciences, 2008

<p>The Arbuscular mycorrhiza (AM) is an endosymbiont of higher plant roots. Most land plants and cultivated crops are concerned to AM symbiosis. This endosymbiosis is based on the mutual exchange of nutrients between plant and fungus. Therefore, AM symbiosis leads to an increased demand for photosynthetic products. The aim of this study was to investigate the pathway used by plants during AM symbiosis to increase photosynthetic performance. Therefore, we have carried out a systematic characterization of photosynthesis in Medicago truncatula (M. truncatula), which is a model legume. We observed colonization by the fungus in roots and that AM symbiosis increases the fresh and dry plant biomass. This could be attributed to an increase in both photosynthetic efficiency and capacity in AM plants. Consistent with these observations, AM symbiosis enhanced phosphorus uptake from the soil into roots, stems and leaves, as based on analyses of phosphorus content. Based on equal chl loading, no differences were found regarding D1, Lhcb1 and Lhcb2 protein content in four plant groups. This indicates similar ratio between chl and PSII proteins. Furthermore, AM symbiosis increases the amount of chlorophyll, steady state oxygen evolution activities, maximum quantum yield (Fv/Fm), and photosynthetic electron transport rate (about 5 fold). Nevertheless, photoprotection was not affected by AM symbiosis. We observed an increase in weight of seed/fruit and weight of seed/plant in AM plants (about 2 fold). Based on these results, we propose that AM symbiosis increases both the efficiency and the capacity of photosynthetic apparatus in the M. truncatula.</p>

The Arbuscular mycorrhiza (AM) is an endosymbiont of higher plant roots. Most land plants and cultivated crops are concerned to AM symbiosis. This endosymbiosis is based on the mutual exchange of nutrients between plant and fungus. Therefore, AM symbiosis leads to an increased demand for photosynthetic products. The aim of this study was to investigate the pathway used by plants during AM symbiosis to increase photosynthetic performance. Therefore, we have carried out a systematic characterization of photosynthesis in Medicago truncatula (M. truncatula), which is a model legume. We observed colonization by the fungus in roots and that AM symbiosis increases the fresh and dry plant biomass. This could be attributed to an increase in both photosynthetic efficiency and capacity in AM plants. Consistent with these observations, AM symbiosis enhanced phosphorus uptake from the soil into roots, stems and leaves, as based on analyses of phosphorus content. Based on equal chl loading, no differences were found regarding D1, Lhcb1 and Lhcb2 protein content in four plant groups. This indicates similar ratio between chl and PSII proteins. Furthermore, AM symbiosis increases the amount of chlorophyll, steady state oxygen evolution activities, maximum quantum yield (Fv/Fm), and photosynthetic electron transport rate (about 5 fold). Nevertheless, photoprotection was not affected by AM symbiosis. We observed an increase in weight of seed/fruit and weight of seed/plant in AM plants (about 2 fold). Based on these results, we propose that AM symbiosis increases both the efficiency and the capacity of photosynthetic apparatus in the M. truncatula.