Final Activity Report Summary - ABC-MEDICAGO (Involvement of ATP binding cassette (ABC) transporters in the plant-microbe interactions in the legume Medicago truncatula)
ATP binding cassette (ABC) proteins form the largest protein family known with more than 1000 members operating in all living cells from bacteria to man. A major function fulfilled by these enzymes is membrane translocation of a wide range of unrelated molecules. Importers constitute mainly the prokaryotic members of the ABC family and are a part of uptake systems for nutrients like amino acids, vitamins, sugars or metal ions. Exporters are found in both prokaryotes and eukaryotes and are involved in extrusion of signalling molecules, toxic substances as well as targeting of membrane components.
Plants are particularly rich in ABC proteins. Arabidopsis has the largest ABC family reported to date with 131 members identified and classified based on their domain organisation (Sanchez Fernandez et al., 2001). ABC transporters were also classified in crops such as rice (Jasinski et al., 2003). To date, most of the ABC proteins that have been characterised in plants are implicated in transport across biological membranes various exogenous (e.g. herbicides, xenobiotics) and endogenous (e.g. phytoalexins, malonylated chlorophyll catabolites, phytohormones, secondary metabolites) compounds.
The observation that different elicitors (bacterial and fungal) trigger the expression of some plant ABC transporters involved in the transport of endogenous defensive compounds led to the conclusion that ABC proteins might play a role in the cross talk between plants and other organisms.
A particular example of plant-microbe interactions is the symbiosis of leguminous plants and endosymbiotic prokaryotes known as rhizobia (Rhizobium, Sinorhizobium). These prokaryotes convert atmospheric N2 to ammonia which is transferred to the plant and incorporated into amino acids. As a consequence, legume plants produce protein-rich seeds and leaves, which are a source of high quality proteins for animal and human nutrition. The symbiosis and formation of a new organ, root nodule, results from the cross talk between plant and rhizobia. Production of lipochitooligosaccharide nodulation factor by rhizobia in response to plant flavonoids secreted into the soil plays an important role in the symbiosis process. Certain secondary metabolites from a model legume plant Medicago, like flavonoid luteolin, which triggers expression of Rhizobium nodulation genes, are substrates for plant ABC transporters. However, in spite of growing amount of biochemical evidence of a potential role of ABC transporters in plant-microbe interaction; little is known about these proteins in leguminous plants.
The specific aim of our study is to determine whether the Medicago ABC transporters are essential components in the cross-talk between plant and rhizobia. We identified in different databases (cDNA libraries derived from elicited Medicago cell cultures or nodulated roots) several ESTs designated as ABC transporters. Using quantitative reverse transcription and PCR (RT-PCR) and real time PCR (Rt-PCR) we have determined the expression pattern of some of them in the roots of Medicago. The complete sequence of a new ABC transporter belonging to the PDR subfamily has been obtained (MtPDR1).
The Rt-PCR transcript analysis of MtPDR1 revealed induction of the gene in the presence of symbiotic bacteria. The promoter sequence of the MtPDR1 gene has been isolated and currently detailed analysis of transcription pattern using GUS reporter system is in progress. The MtPDR1 is also expressed in the suspension cell cultures derived from roots of Medicago. We are using these cultures for phytochemical studies of the changes in the profile of secondary metabolites as a consequence of symbiotic interaction. Possessing the complete sequence of MtPDR1 allows us to design specific antibodies and investigate its expression at the protein level. Our data provide first insights into the ABC transporters from leguminous plants.
Plants are particularly rich in ABC proteins. Arabidopsis has the largest ABC family reported to date with 131 members identified and classified based on their domain organisation (Sanchez Fernandez et al., 2001). ABC transporters were also classified in crops such as rice (Jasinski et al., 2003). To date, most of the ABC proteins that have been characterised in plants are implicated in transport across biological membranes various exogenous (e.g. herbicides, xenobiotics) and endogenous (e.g. phytoalexins, malonylated chlorophyll catabolites, phytohormones, secondary metabolites) compounds.
The observation that different elicitors (bacterial and fungal) trigger the expression of some plant ABC transporters involved in the transport of endogenous defensive compounds led to the conclusion that ABC proteins might play a role in the cross talk between plants and other organisms.
A particular example of plant-microbe interactions is the symbiosis of leguminous plants and endosymbiotic prokaryotes known as rhizobia (Rhizobium, Sinorhizobium). These prokaryotes convert atmospheric N2 to ammonia which is transferred to the plant and incorporated into amino acids. As a consequence, legume plants produce protein-rich seeds and leaves, which are a source of high quality proteins for animal and human nutrition. The symbiosis and formation of a new organ, root nodule, results from the cross talk between plant and rhizobia. Production of lipochitooligosaccharide nodulation factor by rhizobia in response to plant flavonoids secreted into the soil plays an important role in the symbiosis process. Certain secondary metabolites from a model legume plant Medicago, like flavonoid luteolin, which triggers expression of Rhizobium nodulation genes, are substrates for plant ABC transporters. However, in spite of growing amount of biochemical evidence of a potential role of ABC transporters in plant-microbe interaction; little is known about these proteins in leguminous plants.
The specific aim of our study is to determine whether the Medicago ABC transporters are essential components in the cross-talk between plant and rhizobia. We identified in different databases (cDNA libraries derived from elicited Medicago cell cultures or nodulated roots) several ESTs designated as ABC transporters. Using quantitative reverse transcription and PCR (RT-PCR) and real time PCR (Rt-PCR) we have determined the expression pattern of some of them in the roots of Medicago. The complete sequence of a new ABC transporter belonging to the PDR subfamily has been obtained (MtPDR1).
The Rt-PCR transcript analysis of MtPDR1 revealed induction of the gene in the presence of symbiotic bacteria. The promoter sequence of the MtPDR1 gene has been isolated and currently detailed analysis of transcription pattern using GUS reporter system is in progress. The MtPDR1 is also expressed in the suspension cell cultures derived from roots of Medicago. We are using these cultures for phytochemical studies of the changes in the profile of secondary metabolites as a consequence of symbiotic interaction. Possessing the complete sequence of MtPDR1 allows us to design specific antibodies and investigate its expression at the protein level. Our data provide first insights into the ABC transporters from leguminous plants.