Mechanisms of ozone tolerance in rice: characterization of two QTLs affecting leaf bronzing by gene expression profiling and biochemical analyses

Abstract

High surface ozone concentration is increasingly being recognized as a factor that negatively affects crop yields in Asia. However, little progress has been made in developing ozone-tolerant genotypes of rice-Asia's major staple crop. This study aimed to identify possible tolerance mechanisms by characterizing two quantitative trait loci (QTLs) that were previously shown to influence visible leaf symptoms under ozone exposure (120 nl l(-1), 7 h d(-1), 13 d). Two chromosome segment substitution lines (SL15 and SL41) that carried introgressions of the QTLs OzT3 and OzT9, respectively, were exposed to ozone at 120 nl l(-1) along with their parent Nipponbare. In accordance with the expected QTL effect, SL15 showed stronger visible symptoms of ozone damage than Nipponbare, whereas SL41 had fewer symptoms. Gene expression profiling by microarray hybridization yielded 470 probes that were differentially expressed in SL15 and 314 in SL41. Potential tolerance mechanisms were evaluated by investigating changes in gene expression in three general categories. (i) Processes involved in programmed cell death, in which a number of genes related to ethylene or jasmonic acid metabolism or general disease resistance were identified that were differentially regulated in one of the substitution lines. (ii) Biosynthesis of antioxidants. Testing this hypothesis did not reveal any genes differentially regulated between genotypes, and it was thus rejected. (iii) Turnover of antioxidants and enzymatic detoxification of radical oxygen species (ROS), in which a number of differentially regulated genes were also identified. Genes encoding antioxidant enzymes (catalase and peroxidases) tended to be more strongly expressed in SL15. A potential tolerance gene which encodes a putative ascorbate oxidase was identified within the QTL introgression in SL41. This gene showed consistently lower expression in SL41 under ozone exposure across different points in time within independent experiments. Its expression may be involved in mechanisms leading to enhanced ascorbic acid status in SL41 under ozone exposure, and may be linked to a higher concentration of total apoplastic ascorbic acid in SL41 that was observed in an independent experiment.

A Novel Gene, OZONE-RESPONSIVE APOPLASTIC PROTEIN1, Enhances Cell Death in Ozone Stress in Rice

“…A proposed candidate gene, which was annotated as an ASCORBATE OXIDASE (AO; Rice Annotation Project Database [RAP-DB] identifier Os09g0365900), was located near the OzT9 locus and was one of the most highly induced genes in ozone stress in the whole array. The induction of gene expression in ozone stress was significantly lower in the tolerant SL41 than in cv Nipponbare (Frei et al, 2010). From a physiological point of view, apoplastic ascorbate (AsA) has been shown to form the first line of defense against the ozone entering the plants through the stomata (Luwe et al, 1993;Plöchl et al, 2000;Feng et al, 2010), which supports the link between the annotation of the candidate gene and ozone stress tolerance.…”

“…We also analyzed the OsORAP1 sequences in a previously reported ozone-tolerant chromosome segment substitution line, SL41, which was genetically identical to cv Nipponbare except for a 13-Mb introgression including the OzT9 region and another introgression on chromosome 6 from the tolerant cv Kasalath (Frei et al, 2010). Our analysis showed that the sequence of SL41 at the OsORAP1 locus was identical to that of cv Kasalath (Supplemental Fig.…”

“…In this study, we characterized OsORAP1, which emerged as a candidate gene for ozone stress tolerance (i.e. the formation of cell death) in a previous study (Frei et al, 2010). To elucidate the physiological function of OsORAP1 in ozone stress, three hypotheses were tested.…”

“…Underpinning the effect of OzT9, a chromosomal segment substitution line, SL41, carrying a chromosomal introgression from cv Kasalath at the OzT9 locus in the cv Nipponbare background, produced less visible symptoms than cv Nipponbare (Frei et al, 2008). A subsequent transcriptomic analysis revealed a differential gene expression profile between cv Nipponbare and SL41 (Frei et al, 2010). A proposed candidate gene, which was annotated as an ASCORBATE OXIDASE (AO; Rice Annotation Project Database [RAP-DB] identifier Os09g0365900), was located near the OzT9 locus and was one of the most highly induced genes in ozone stress in the whole array.…”

Emission inventories and projections

“…Despite the overwhelming evidence that current O 3 concentrations are causing yield losses, new O 3 tolerant crop cultivars are not being developed for a future higher-O 3 world [Ainsworth et al, 2008;Booker et al, 2009]. Recent successes in identifying quality trait loci associated with O 3 tolerance in rice indicate the breeding for O 3 tolerance in food crops is possible [Frei et al, 2008;Frei et al, 2010], yet currently, there is little if any industrial effort in this direction.…”