| One of the most remarkable achievements in nature is the evolution of the semiautonomous organelles including chloroplasts and mitochondria in eukaryotic cells via endosymbiosis. Chloroplast development requires the coordinated expressions of nuclear and chloroplast genomes, and both anterograde and retrograde signals exist and work together to facilitate this coordination.Arabidopsis yellow variegated2(var2) mutant was used as an entry point to dissect the genetic regulatory network of chloroplast development. Here, I report the isolation of two new var2 genetic suppresors, svr9-1(suppressor of variegation 9-1) and svr11-1(suppressor of variegation 11-1). Based on gene cloning, gene functional study, protein subcellular localization analysis, tissue expression assay, var2 variegation inhibition mechanism study and leaf morphology regulation analysis to explore the roles of SVR9, SVR9-Like1(SVR9L1) and SVR11 play in chloroplast and plant development1. The results about SVR9 and SVR9L1 are as follows:(1) Arabidopsis thaliana svr9-1 mutant was isolated as var2-5 suppressor via activation tagging method. SVR9 gene(At2g24060) encoding a chloroplast translation initiation factor IF3 was identified by map based cloning and complementation assay. svr9-1 mutant can be fully rescued by the Escherichia coli IF3 infC, suggesting that SVR9 functions as a bona fide IF3 in the chloroplast.(2) SVR9 and its close homologue SVR9L1 both targeted to chloroplast. Constitutive expression of SVR9L1 rescue svr9-1 phenotype demonstrates that SVR9 and SVR9L1 are functionaly redundant. Homozygous svr9-1 svr9l1-1 mutant was not able to get through genetic cross experiment, while the mutant homozygous for svr9-1 but heterozygous for svr9l1-1(svr9-1 svr9l1-1/+) was recovered. It indicates that functional IF3 s are essential to plant survival and development.(3) Down-regulation of chloroplast IF3s' activities not only caused chloroplast development defects, but also led to a series of leaf developmental abnormalities including more serrated leaf margin, disorganized mesophyll cells and altered cotyledon venation patterns. Moreover, auxin homeostasis is disturbed both in chloroplast IF3 s defective mutants and in plants treated with chloroplast translation inhibitors.(4) Genetic analysis between SVR9, SVR9L1 and VAR2 indicate that chloroplast developmental state impact VAR2 threshold. Genetic analysis established that SVR9/SVR9L1-mediated leaf margin development is dependent on CUC2 activities and is independent of their roles in chloroplast development. That is, chloroplast translation initiation factors SVR9 and SVR9L1 are required not only for chloroplast development and var2-mediated leaf variegation, but also are involved in the coordination of leaf and chloroplast development.2. The results about SVR11 are as follows:(1) svr11-1 mutant with a virescent leaf phenotype was isolated as var2-5 suppressor via activation tagging method. SVR11(At4g20360) encodes a chloroplast translation elongation factor EF-Tu was determined by inverse PCR and complementation analysis. SVR11 targeted to chloroplast and SVR11 tissue expression is regulated by plant development.(2) Genetic analysis through three different allels of SVR11(svr11-1, svr11-2 and svr11-3) shows decreased expression of SVR11 results in virescent, yellow-colored leaves with lobation, enventurally embryo lethal phenotypes. SVR11 activity is required for plant development.(3) Abnormalities in leaf anatomy, cotyledon venation patterns and leaf margin development were identified in svr11-1 svr9-1 double mutant further support that chloroplast states impact leaf development.From all above, we conclude that chloroplast translation initiation factor IF3 s and elongation factor EF-Tu are indispensable regulators in chloroplast and leaf development. |
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