The programme is aimed at the identification of several membrane proteins directly involved in the distribution of assimilates and ions in plants, and at the cloning of the corresponding genes. These proteins include the sucrose carrier, the amino acid carriers and the ion carriers of the plasmalemma, and the malate transporter of the tonoplast.
Crop productivity depends on the ability of the plant to transport the assimilates from the leaf blade to the receiving organs harvested for human consumption. This transport is controlled by the distribution and the activity of specific membrane transporters. The objective of the project is to characterize and identify plant plasmalemma transporters and tonoplast transporters, and to clone the corresponding genes. Sucrose is the major mobile carbohydrate in the plant and may be finally stored as sucrose or as starch in fruits. Amino acid transport is necessary for the synthesis of storage proteins found in cereals and legumes. Malate is one of the major solutes accumulated in vacuoles of higher plants, being involved in transient storage of carbon, charge balance, cytoplasmic homeostasis and turgor maintenance. Ion transport across the plasmalemma is necessary for the mainteance of cell metabolism.
Significant work to date includes:
reconstitution procedures for the sucrose carrier and amino acid carriers;
complementation studies with yeasts;
the first demonstration that a plant membrane protein can be successfully synthesized and targeted to the oocyte plasma membrane;
isolation of Arabidopsis mutants for K plus uptake;
identification of a novel malate selective ion channel in the tonoplast of CAM plants;
establishment of a library of monoclonal antibodies against the native tonoplast membrane of Kalancthoe;
development of a protocol for isolation of ribonucleic acid (RNA) and deoxyribonucleic acid (DNA) from Kalanchoe mesophyll cells.
From current progress it may be foreseen that geneticengineering of transport proteins in plants will be possible in the near future. This opens new insights on possible manipulation of the plant content and on its stress tolerance.
Plant productivity is dependent on numerous events mediated by membrane proteins specialized in the transport of organic solutes (sucrose, amino acids, malate, and of ions (H+, K+). Indeed long distance transport of assimilate from the source leaf to the sink org.(tubers, roots, fruits) which are harvested is controlled by the transport processes occurring across the plasmalemma and across the tonoplast of the leaf cells (source) and of the storage cells (sink). The Proposal intends (1) to identify the Proteins involved in the transport of sucrose, of amino acids, and of K+ across the plasmalemma (2) to identify the proteins involved in the transport of malate across the tonoplast (3) to isolate and to clone the genes cod for the sucrose carrier and for the malate carrier (4) to study the expression of the gene coding for the sucrose carrier in various tissues and various organs, after fusion with different promoters already available (5) to study the targeting of the sucrose carrier to the plasmal. or to the tonoplast. Previous work published by one of the groups involved in the project has identified a good candidate for the sucrose carrier of the plasmalemma, by means of differential labeling and by use of polyclonal antibodies. Monoclonal antibodies are presently being prepared, and the general expertise of the group will be used to develop similar Procedures for the amino acid car.of the Plasmalemma. The malate carrier will be identified by two groups already working on this topic. Mutants of Arabidopsis deficient in ion transport will be selected by another group, with the help of a private subcontractor. At last, two groups internationally recognized for their experience in molecular biology will develop this a.s.a.p. especially on the sucrose carrier.
Funding SchemeCSC - Cost-sharing contracts
43036 Fidenza Parma
AL5 2JQ Harpenden, Herts
OX1 3RB Oxford