Fine Mapping And Cloning Of Gene PSLS1 In Rice(Oryza Sativa L.)

As we all known, blade is an important source organ for plants. During leaf senescence in rice,chlorophyll, proteins and other macromolecules are degraded, reducing leaf photosynthesis and following with accumulation of ROS and increasing of SAGs expression. All of these accelerate cell death and decrease crop yield and grain quality. Hence, it is of great importance to identify novel mutants and characterize their molecular and physiological mechanisms. In this study, we reported the gene mapping and cloning of a psls1 mutant(premature senescence leaf with spots) in rice. We found that a single-base substitution of PSLS1, encoding a ferredoxin-dependent glutamate synthase(Fd-GOGAT), leaded to early senescence in psls1.Main results as follows:1. The mutant showed early senescence symptoms from bottom to top of plants after 7-leaf stage and the chlorisis firstly appeared in leaf opex and then spread to whole leaf with brown spots. What’s worse, psls1 overall withered at maturity stage. Compared with wide type, the plant height and panicle length shortened, the number of tiller and spikelets per panicle were significantly decreased, 1000-grain weight reduced, and the seeding rate also declined slightly.2. There were significantly physiological difference between wide type and psls1, including decreased chlorophyll content, over-accumulated H2O2, yellow-withered leaf and increased dead cell from bottom to top of plant after the 7-leaf stage. Observation by transmission electron microscope(TEM) showed degradation of chloroplast, detective thylakoid and numerous osmiophilic granules in psls1 leaves.3. Genetic analysis demonstrated that the phenotype of psls1 was caused by a single recessive gene.Using genetic population derived from psls1 × IRAT129, the psls1 locus was mapped in a 89 kb region flanked by markers zs-3 and zs-8 on chromosome 7. This region contains 12 putative open reading frames(ORFs), sequence analysis revealed that G was replaced with A in the junction of the second exon and intron of LOC_Os07g46460, which lead to a 59 bp deletion in c DNA. Therefore,LOC_Os07g46460, encoding a ferredoxin-dependent glutamate synthase, predicted to be the candidate gene and was designated as PSLS1.4. Consistent with previous reports,PSLS1 was constitutively expressed in all tissues, and had the highest expression level in leaves. There was no significantly affected of m RNA expression level of PSLS1 in the psls1 seedlings, but it decreased sharply after chlorisis. Moreover, the suppression of PSLS1 transcripts resulted in reduction of glutamate synthase activity(GOGAT).5. In heading date, the content of free amino acids in psls1 was clearly different from that in wide type. For example Glu, Ser, Gly, Ala, Val and Ile all dropped obviously, whereas, the contents of Asn,PSer, Thr, Cys, Phe, His, Orn, Lys and Arg increased significantly. These indicated abnormal amino acid metabolism in psls1. Under low nitrogen treatment, the senescence phenotype of psls1 could occur as early as at 3-leaf stage. These results indicated that psls1 senescence phenotype might be associated with the loss of glutamate synthase activity and the abnormal amino acid metabolism.