Poly(ADP-ribose)polymerase activity controls plant growth by promoting leaf cell number

A changing global environment, rising population and increasing demand for biofuels are challenging agriculture and creating a need for technologies to increase biomass production. Here we demonstrate that the inhibition of poly (ADPribose) polymerase activity is a promising technology to achieve this under non-stress conditions. Furthermore, we investigate the basis of this growth enhancement via leaf series and kinematic cell analysis as well as single leaf transcriptomics and plant metabolomics under non-stress conditions. These data indicate a regulatory function of PARP within cell growth and potentially development. PARP inhibition enhances growth of Arabidopsis thaliana by enhancing the cell number. Time course single leaf transcriptomics shows that PARP inhibition regulates a small subset of genes which are related to growth promotion, cell cycle and the control of metabolism. This is supported by metabolite analysis showing overall changes in primary and particularly secondary metabolism. Taken together the results indicate a versatile function of PARP beyond its previously reported roles in controlling plant stress tolerance and thus can be a useful target for enhancing biomass production

Mapping the Arabidopsis Metabolic Landscape by Untargeted Metabolomics at Different Environmental Conditions

Metabolic genome-wide association studies (mGWAS), whereupon metabolite levels are regarded as traits, can help unravel the genetic basis of metabolic networks. A total of 309 Arabidopsis accessions were grown under two independent environmental conditions (control and stress) and subjected to untargeted LC-MS-based metabolomic profiling; levels of the obtained hydrophilic metabolites were used in GWAS. Our two-condition-based GWAS for more than 3000 semi-polar metabolites resulted in the detection of 123 highly resolved metabolite quantitative trait loci (p ≤ 1.0E-08), 24.39% of which were environment-specific. Interestingly, differently from natural variation in Arabidopsis primary metabolites, which tends to be controlled by a large number of small-effect loci, we found several major large-effect loci alongside a vast number of small-effect loci controlling variation of secondary metabolites. The two-condition-based GWAS was followed by integration with network-derived metabolite-transcript correlations using a time-course stress experiment. Through this integrative approach, we selected 70 key candidate associations between structural genes and metabolites, and experimentally validated eight novel associations, two of them showing differential genetic regulation in the two environments studied. We demonstrate the power of combining large-scale untargeted metabolomics-based GWAS with time-course-derived networks both performed under different abiotic environments for identifying metabolite-gene associations, providing novel global insights into the metabolic landscape of Arabidopsis. By combining large-scale untargeted metabolomics-based GWAS and network analysis with environmental stress-driven perturbations of metabolic homeostasis, this system-wide study provides new global insights into the metabolic landscape of Arabidopsis, using a strategy that could readily be extended to other plant species.</p

Ploidy analysis of leaf two.

<p>Arabidopsis leaf two was analyzed at the indicated time points from plants subjected to PARP inhibition or control conditions. (A) the calculated EI is shown, EI represents the number of endo cycles undergone by a typical nuclei (EI  =  1*4N+2*8N+3*16N), in (B–D) percentage of 2N, 4N and 8N nuclei is presented. 10–12 leaf two were pooled for a replicate, four biological replicates were analyzed in each of the two independent experiments (n = 8).</p

PARP inhibition altering secondary metabolism.

<p>Arabidopsis shoots were harvested at day7 after transfer. Per replicate 10–12 seedlings were harvested, with 5 biological replicates in each of the three independent experiments (n = 15) and analyzed using LC-MS. The five strongest up- and down-regulated metabolites are shown.</p

PARP activity and effects on primary metabolism.

<p>The metabolite profile of Arabidopsis shoots was analyzed by GC-MS at the indicated times following transfer of the seedlings after 7 days growth on control media to either control or PARP inhibitor containing media. Plants were grown in parallel to those for the physiological and microarray analysis. Five individual samples, each a pool of 10–12 plants, were harvested and analyzed in three independent experiments (n = 15). Amino, organic and fatty acids with significant changes in their relative abundance are shown. Metabolite content relative to the (−3MB), with red and blue indicating accumulation and depletion, are shown.</p

PARP inhibition and its effect on guard cell development.

<p>Arabidopsis leaf two was analyzed after two days of transferring the seedlings (day2) or after seven days (day7) either to treatment or control conditions. (A) The stomata index (SI) is shown, calculated by number of stomata divided by number of cells, (B) the number of stomata per mm² and (C) the calculated total number of stomata per leaf is presented. In each experiment 4–6 leafs were analyzed and the experiment repeated three times independently (n = 12–18). Significant differences (P<0.05) between treated and untreated plants is indicated by an asterisk.</p

PARP inhibition and its effect on the phenylpropanoid pathway.

<p>Arabidopsis shoots were harvested at day7 after transfer. Per replicate 10–12 seedlings were harvested, with 5 biological replicates in each of the three independent experiments (n = 15). The metabolite content relative to the control (no 3MB) is shown and when changed significantly the relative content is underlined. Only metabolites with a clear annotation and positioning within the pathway are shown.</p