Functional Identification Of Two Tomoto Phosphater Transporter Genes LePT1 And LePT2 In Rice

Phosphorus (P) is one of the essential macronutrients for plant growth and development. It plays key roles in plant metabolic processes, such as energy transfer, signal transduction, biosynthesis of macromolecules. The primary source for P uptake by transporters of plant roots is inorganic phosphate (Pi) in soils. Pi mobilization is very low in soils due to microbial and metal ion fixation. Plants have evolved several strategies to improve Pi acquisition, including alterations in root morphology, modification of soil chemical properties around the roots, formation of symbiotic associations with arbuscular mycorrhizal (AM) fungi and activation of high-affinity Pi transporters.Molecular breeding of high Pi use efficient crops by exploitation and transformation of the key genes regulating Pi acquisition and utilization has become one of the hotest research topics in current plant biology. It is known that plants use specific Pi transporters (Pht) in the roots to take up Pi from soil solution and Pi translocation within the plants. So far, most of the identified Pht members in plants belong to putatively high affinity Pht1 family. Previous researches had shown that LePT1 and LePT2 are two tomato Phtl genes regulated by Pi-starvation.In this study, we first cloned LePT1 and LePT2 from tomato genome, then expressed them in a rice cultivar (Nipponbare) through the transgenic technology with the Agrobacterium tumefaciens. The bio-informatics of the gene sequences and physiological analyses of the transgenic rice showed the following results:1. The cluster analysis of the predicted amino acid sequences and transmembrane domains demonstrated that LePT1 and LePT2 are highly conserved in the structure and functional site with 13 members of Pht1 family in rice genome. They show the highest similarity with Pi-starvation regulated rice Pi transporters, OsPT2 and OsPT4, respectively. 2. Semi-quantitative RT-PCR analyses indicated that the respective abundant transcript levels of tomato LePT1 and LePT2 driven by the constitutive promoter CaMV35S in both roots and shoots of the individual transgenic lines of rice. Interestingly, the expression of either LePT1 or LePT2 did not affect the transcript levels of each endogenous Phtl members in the rice.3. Four independent transgenic lines of rice with expression of LePTl and LePT2 were selected for analyzing their responses to Pi starvation and sufficient supply in comparison to the non transgenic rice (WT). Expression of either LePT1 or LePT2 in the rice resulted significant increases of water extractable and total P concentrations, plant height and biomass yield at low Pi supply condition. However, the transgenic rice accumulated very high P concentrations in comparison to WT at high Pi supply condition, which might be the cause of relative smaller size and less tiller number by the constitutive expression of LePT1 and OsPT2.Taken together, the resulted confirmed that both LePTl and LePT2 have the functions of Pi uptake and translocation, the Phtl members from di-cotyledons could be directly used for improving Pi use in mono-cotyledons through molecular genetics approach.