Dissecting rice polyamine metabolism under controlled long-term drought stress

Abstract

A selection of 21 rice cultivars (Oryza sativa L. ssp. indica and japonica) was characterized under moderate long-term drought stress by comprehensive physiological analyses and determination of the contents of polyamines and selected metabolites directly related to polyamine metabolism. To investigate the potential regulation of polyamine biosynthesis at the transcriptional level, the expression of 21 genes encoding enzymes involved in these pathways were analyzed by qRT-PCR. Analysis of the genomic loci revealed that 11 of these genes were located in drought-related QTL regions, in agreement with a proposed role of polyamine metabolism in rice drought tolerance. The cultivars differed widely in their drought tolerance and parameters such as biomass and photosynthetic quantum yield were significantly affected by drought treatment. Under optimal irrigation free putrescine was the predominant polyamine followed by free spermidine and spermine. When exposed to drought putrescine levels decreased markedly and spermine became predominant in all cultivars. There were no correlations between polyamine contents and drought tolerance. GC-MS analysis revealed drought-induced changes of the levels of ornithine/arginine (substrate), substrates of polyamine synthesis, proline, product of a competing pathway and GABA, a potential degradation product. Gene expression analysis indicated that ADC-dependent polyamine biosynthesis responded much more strongly to drought than the ODC-dependent pathway. Nevertheless the fold change in transcript abundance of ODC1 under drought stress was linearly correlated with the drought tolerance of the cultivars. Combining metabolite and gene expression data, we propose a model of the coordinate adjustment of polyamine biosynthesis for the accumulation of spermine under drought conditions.

Figure 1. Mean rank of stress damage evaluated by visual scoring after 18 days of drought treatment.

Each value represents the mean rank (±SE) of three experiments with five replicates each. Score of control plants was 1 (no damage) for 94% of the plants. Cultivars are ordered by mean rank. For cultivar numbers see Table 1.

Figure 2. Polyamine content under control and drought conditions.

The different panels show Put (A), Spd (B) and Spm (C) content of leaves of 21 rice cultivars. Cultivars numbered from 1 to 50 were sorted from most tolerant to most sensitive. Each value represents the mean (±SE) of two experiments with five replicates each.

Figure 3. Fold change of selected metabolites under drought in comparison to control conditions.

Fold change (log2) of metabolites in leaves of 21 cultivars is shown. Cultivars numbered from 1 to 50 were sorted from most tolerant to most sensitive. Data represent the means of five biological replicates from one experiment.

Figure 4. Expression of genes encoding enzymes involved in Put biosynthesis under control and drought conditions.

Panel (A) shows relative gene expression (log2) under control conditions determined for nine selected cultivars. Panel (B) shows the log2 fold change values for the gene expression under drought compared to control conditions. Data represent the means of two biological experiments with three technical replicates each. Cultivars were sorted from most tolerant to most sensitive.

Figure 5. Expression of genes encoding enzymes involved in Spd/Spm biosynthesis under control and drought conditions.

Panel (A) shows relative gene expression (log2) under control conditions determined for nine selected cultivars. Panel (B) shows the log2 fold change values for the gene expression under drought compared to control conditions. Data represent the means of two biological experiments with three technical replicates each. Cultivars were sorted from most tolerant to most sensitive.

Figure 6. Model for the regulation of polyamine metabolism in response to drought stress in rice leaves.

Red - decrease, blue – increase, green – cultivar dependent, black – not measured. Arg, arginine; Orn, ornithine; SAM, S-adenosylmethionine; dcSAM, decarboxylated S-adenosylmethionine.