Cellular Localization And Functional Analysis Of FDR3 And FDR4

fdr (Fe-deficiency-related) is one of the differential screened clones from cDNA expression library of Fe-deficient-induced maize root. The fdr with the full length of 3.3kb fragment contains a 1068bp ORF1 and a 648bp ORF2. Compared homologous of fdr with other gene, no any homologous gene was found with known genes until now. fdr3 and fdr4, at the front of which is the his₆-c-myc epitope, have been cloned into plant expression vector pBI121 to construct recombinants T-DNA-his₆-c-myc::fdr3(Fdr3 for short) and T-DNA-his₆-c-myc::fdr4 (Fdr4 for short) respectively.I transfered the plant expression vectors Fdr3 and Fdr4 into Agrobacterium EHA105 and then transformed it into the wild tobacco SRI by Agrobacterium- mediated of leaf disc method. After that, we gained some Kanamycin-resistant transgenic tobaccos, which were identified by the molecular biology protocols, such as PCR, RT-PCR, and so on. At last , T₀ seeds were grown in the soil, and T₁ seeds were gained .I identified T₁ tobaccos in the molecular level, and then, growed the transgenic ones which were identified as positive clone in four kinds of liquid iron-nutritional culture medium. It had not been found that there were obvious differences in the leaves' or roots' phenotype, but leaves were different in green gradient. Maybe the chlorophyll content in the leaves was not the same.Using transmission electron microscope, I had found that there were obvious differences in the content of chloroplast and amyloplast and in the structure of grana lamella between transformed and wild tobaccos. Two proteins MYC-FDR3 and MYC-FDR4 were found to be localized in the chloroplast with fluorescent immunocytochemistry. With the bioinformatics method, I infered that FDR3 protein might be localized in the chloroplast thylakoid, and FDR4 protein might be localized on the membrane; FDR3 protein would be a soluble protein without a transmembrane domain or signal peptide, and it contains the similar domain with FliN-protein's; FDR4 protein would be a secretory protein with four transmembrane domains and signal peptide, it contains the similar domain with FliP-protein's. They all have some protein kinase phosphorylation sites which are related to signal transduction.It was persumed that the protein FDR3 and FDR4 might be the components in the protein transport channel on the thylakoid membrane. FDR3 protein might be a switch component to the transport channel of external proteins entering into membrane, whose function was similar to the FliN's. The function of FDR4 protein might be related to protein transport. It's surmised that the genes which encoded the protein of FDR3 and FDR4 might be integrated to the plant genome though symbiosis and be kept down with longtime evolution after the original prokaryote bacteria infecting into the plant.