Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of Arabidopsis shoots from plants grown with low Molybdenum (Mo)

ABSTRACT: ABSTRACT; Molybdenum (Mo) is an essential micronutrient for plants, serving as a cofactor for enzymes involved in nitrate assimilation, sulfite detoxification, abscisic acid biosynthesis and purine degradation. Here we show that natural variation in shoot Mo content across 92 Arabidopsis thaliana accessions is controlled by variation in a mitochondrially localized transporter (Molybdenum Transporter 1 -MOT1) that belongs to the sulfate transporter superfamily. A deletion in the MOT1 promoter is strongly associated with low shoot Mo, occurring in seven of the accessions with the lowest shoot content of Mo. Consistent with the low Mo phenotype, MOT1 expression in low Mo accessions is reduced. Reciprocal grafting experiments demonstrate that the roots of Ler-0 are responsible for the low Mo accumulation in shoot, and GUS localization demonstrates that MOT1 is expressed strongly in the roots. MOT1 contains an N-terminal mitochondrial targeting sequence, and expression of MOT1 tagged with GFP in protoplasts, and transgenic plants, establishes the mitochondrial localization of this protein. Furthermore, expression of MOT1 specifically enhances Mo accumulation in yeast by 5-fold, consistent with MOT1 functioning as a molybdate transporter. This work provides the first molecular insight into the processes that regulate Mo accumulation in plants, and shows that novel loci can be detected by association mapping. Experiment Overall Design: Hybridizations from a set of Bulk Segregant analysis. An F2 population from 14501 in the col-0 background crossed to Ler-0 was analyzed. This series contains the 3 hybs from Ler used to identify Single Feature Polymorphisms, the 3 hybs of Col-0 they were compared to, and 1 hyb for each pool from the BSA mapping(High Mo pool, low Mo Pool).

ORGANISM(S): Arabidopsis thaliana

SUBMITTER: Ivan Baxter

PROVIDER: E-GEOD-10039 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Background:; Plants are sessile and therefore have developed mechanisms to adapt to their environment, including the soil mineral nutrient composition. Ionomics is a developing functional genomics strategy designed to rapidly identify the genes and gene networks involved in regulating how plants acquire and accumulate these mineral nutrients from the soil. Here we report on the coupling of high-throughput elemental profiling of shoot tissue from various Arabidopsis accessions with DNA microarray-based bulk segregant analysis (BSA) and reverse genetics, for the rapid identification of genes from wild populations of Arabidopsis that are involved in regulating how plants acquire and accumulate Na+ from the soil. Methodology/Principal Findings:; Elemental profiling of shoot tissue from 12 different Arabidopsis accessions revealed that Ts-1 and Tsu-1 accumulate higher shoot levels of Na+ than Col-0 and other accessions. We identify AtHKT1, known to encode a Na+ transporter, as being the causal locus driving elevated shoot Na+ in both Ts-1 and Tsu-1. Furthermore, we establish that a deletion in a tandem repeat sequence ~5 kb upstream of AtHKT1 is responsible for the reduced root expression of AtHKT1 observed in these accessions. Reciprocal grafting experiments establish that this loss of AtHKT1 expression in roots is responsible for elevated shoot Na+. Interestingly, and in contrast to the hkt1-1 null mutant, under NaCl stress conditions this novel AtHKT1 allele not only does not confer NaCl sensitivity, but co-segregates with elevated NaCl tolerance. We also present all our elemental profiling data in a new open access ionomics database, the Purdue Ionomics Information Management System (PiiMS; www.purdue.edu/dp/ionomics). Conclusions/Significance:; Using DNA microarray-based genotyping has allowed us to rapidly identify AtHKT1 as the causal locus driving the natural variation in shoot Na+ accumulation we observed in Ts-1 and Tsu-1, two coastal populations of Arabidopsis. Such an approach overcomes the limitations imposed by a lack of established genetic markers in most Arabidopsis accessions, and opens up a vast and tractable source of natural variation for the identification of gene function not only in ionomics but also in many other biological processes. Experiment Overall Design: Hybridizations from two sets of Bulk Segregant analysis. F2 populations from col-0 crossed to two High Na accessions: Ts-1 and Tsu-1 were analyzed. This series contains the 3 hybs from each accession that were used to identify Single Feature Polymorphisms, the 3 hybs of Col-0 they were compared to, and 1 hyb for each pool from the BSA mapping(High Na pool, low Na Pool).

Project description:ABSTRACT: Sphingolipid synthesis is initiated by condensation of serine with palmitoyl-CoA to produce 3-ketodihydrosphinganine (3-KDS), which is subsequently reduced by a 3-KDS reductase to dihydrosphinganine (DHS). Serine palmitoyltransferase was recently shown to be essential for plant viability, but the 3-KDS reductase step of sphingolipid synthesis has not been investigated in plants. Arabidopsis thaliana contains two genes (At3g06060/TSC10A and At5g19200/TSC10B) that encode proteins with significant homology to the yeast 3-KDS reductase, Tsc10p. Heterologous expression, in yeast, of either A. thaliana gene restored 3-KDS reductase activity to the yeast tsc10Δ mutant, thereby identifying both as bona fide 3-KDS reductase genes. Consistent with previous evidence that sphingolipids have essential functions in plants, double mutant progeny lacking both genes were not recovered. Although the 3-KDS reductase genes are functionally redundant and ubiquitously expressed in A. thaliana, 3-KDS reductase activity was reduced to approximately 10% of wild-type levels in the loss-of-function tsc10a mutant, leading to an altered sphingolipid profile compared to wild-type plants. Interestingly, this perturbation of sphingolipid biosynthesis in the A. thaliana tsc10a mutant leads to significant alterations in the leaf ionome, including increases in Na, K and Rb (a K analogue), and decreases in Mg, Ca, Fe, and Mo. Reciprocal grafting revealed that these changes in the leaf ionome are driven by the root, and are associated with both increases in root suberin, and elevation of expression in the root of genes involved in Fe homoeostasis, including the Fe-transporter IRT1. Keywords: genomic hybridization bulked segregant analysis Hybridizations from a set of Bulk Segregant analysis. An F2 population from 7113 in the col-0 background crossed to Ler-0 was analyzed. This series contains the 3 hybs from Ler used to identify Single Feature Polymorphisms, the 3 hybs of Col-0 they were compared to, and 1 hyb for each pool from the BSA mapping(low Ca/Mg Pool and wild type pools).

Project description:Background: Plants are sessile and therefore have developed mechanisms to adapt to their environment, including the soil mineral nutrient composition. Ionomics is a developing functional genomics strategy designed to rapidly identify the genes and gene networks involved in regulating how plants acquire and accumulate these mineral nutrients from the soil. Here we report on the coupling of high-throughput elemental profiling of shoot tissue from various Arabidopsis accessions with DNA microarray-based bulk segregant analysis (BSA) and reverse genetics, for the rapid identification of genes from wild populations of Arabidopsis that are involved in regulating how plants acquire and accumulate Na+ from the soil. Methodology/Principal Findings: Elemental profiling of shoot tissue from 12 different Arabidopsis accessions revealed that Ts-1 and Tsu-1 accumulate higher shoot levels of Na+ than Col-0 and other accessions. We identify AtHKT1, known to encode a Na+ transporter, as being the causal locus driving elevated shoot Na+ in both Ts-1 and Tsu-1. Furthermore, we establish that a deletion in a tandem repeat sequence ~5 kb upstream of AtHKT1 is responsible for the reduced root expression of AtHKT1 observed in these accessions. Reciprocal grafting experiments establish that this loss of AtHKT1 expression in roots is responsible for elevated shoot Na+. Interestingly, and in contrast to the hkt1-1 null mutant, under NaCl stress conditions this novel AtHKT1 allele not only does not confer NaCl sensitivity, but co-segregates with elevated NaCl tolerance. We also present all our elemental profiling data in a new open access ionomics database, the Purdue Ionomics Information Management System (PiiMS; www.purdue.edu/dp/ionomics). Conclusions/Significance: Using DNA microarray-based genotyping has allowed us to rapidly identify AtHKT1 as the causal locus driving the natural variation in shoot Na+ accumulation we observed in Ts-1 and Tsu-1, two coastal populations of Arabidopsis. Such an approach overcomes the limitations imposed by a lack of established genetic markers in most Arabidopsis accessions, and opens up a vast and tractable source of natural variation for the identification of gene function not only in ionomics but also in many other biological processes. Keywords: genomics hybridization bulk segregant analysis

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Transcription profiling of Arabidopsis shoots from plants grown with low Molybdenum (Mo)

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