The physiological basis for genetic variation in water use efficiency and carbon isotope composition in Arabidopsis thaliana.

Ecologists and physiologists have documented extensive variation in water use efficiency (WUE) in Arabidopsis thaliana, as well as association of WUE with climatic variation. Here, we demonstrate correlations of whole-plant transpiration efficiency and carbon isotope composition (δ(13)C) among life history classes of A. thaliana. We also use a whole-plant cuvette to examine patterns of co-variation in component traits of WUE and δ(13)C. We find that stomatal conductance (g s) explains more variation in WUE than does A. Overall, there was a strong genetic correlation between A and g s, consistent with selection acting on the ratio of these traits. At a more detailed level, genetic variation in A was due to underlying variation in both maximal rate of carboxylation (V cmax) and maximum electron transport rate (Jmax). We also found strong effects of leaf anatomy, where lines with lower WUE had higher leaf water content (LWC) and specific leaf area (SLA), suggesting a role for mesophyll conductance (g m) in variation of WUE. We hypothesize that this is due to an effect through g m, and test this hypothesis using the abi4 mutant. We show that mutants of ABI4 have higher SLA, LWC, and g m than wild-type, consistent with variation in leaf anatomy causing variation in g m and δ(13)C. These functional data also add further support to the central, integrative role of ABI4 in simultaneously altering ABA sensitivity, sugar signaling, and CO2 assimilation. Together our results highlight the need for a more holistic approach in functional studies, both for more accurate annotation of gene function and to understand co-limitations to plant growth and productivity

Pleiotropy of FRIGIDA enhances the potential for multivariate adaptation.

An evolutionary response to selection requires genetic variation; however, even if it exists, then the genetic details of the variation can constrain adaptation. In the simplest case, unlinked loci and uncorrelated phenotypes respond directly to multivariate selection and permit unrestricted paths to adaptive peaks. By contrast, 'antagonistic' pleiotropic loci may constrain adaptation by affecting variation of many traits and limiting the direction of trait correlations to vectors that are not favoured by selection. However, certain pleiotropic configurations may improve the conditions for adaptive evolution. Here, we present evidence that the Arabidopsis thaliana gene FRI (FRIGIDA) exhibits 'adaptive' pleiotropy, producing trait correlations along an axis that results in two adaptive strategies. Derived, low expression FRI alleles confer a 'drought escape' strategy owing to fast growth, low water use efficiency and early flowering. By contrast, a dehydration avoidance strategy is conferred by the ancestral phenotype of late flowering, slow growth and efficient water use during photosynthesis. The dehydration avoidant phenotype was recovered when genotypes with null FRI alleles were transformed with functional alleles. Our findings indicate that the well-documented effects of FRI on phenology result from differences in physiology, not only a simple developmental switch

Ischaemic accumulation of succinate controls reperfusion injury through mitochondrial ROS.

Ischaemia-reperfusion injury occurs when the blood supply to an organ is disrupted and then restored, and underlies many disorders, notably heart attack and stroke. While reperfusion of ischaemic tissue is essential for survival, it also initiates oxidative damage, cell death and aberrant immune responses through the generation of mitochondrial reactive oxygen species (ROS). Although mitochondrial ROS production in ischaemia reperfusion is established, it has generally been considered a nonspecific response to reperfusion. Here we develop a comparative in vivo metabolomic analysis, and unexpectedly identify widely conserved metabolic pathways responsible for mitochondrial ROS production during ischaemia reperfusion. We show that selective accumulation of the citric acid cycle intermediate succinate is a universal metabolic signature of ischaemia in a range of tissues and is responsible for mitochondrial ROS production during reperfusion. Ischaemic succinate accumulation arises from reversal of succinate dehydrogenase, which in turn is driven by fumarate overflow from purine nucleotide breakdown and partial reversal of the malate/aspartate shuttle. After reperfusion, the accumulated succinate is rapidly re-oxidized by succinate dehydrogenase, driving extensive ROS generation by reverse electron transport at mitochondrial complex I. Decreasing ischaemic succinate accumulation by pharmacological inhibition is sufficient to ameliorate in vivo ischaemia-reperfusion injury in murine models of heart attack and stroke. Thus, we have identified a conserved metabolic response of tissues to ischaemia and reperfusion that unifies many hitherto unconnected aspects of ischaemia-reperfusion injury. Furthermore, these findings reveal a new pathway for metabolic control of ROS production in vivo, while demonstrating that inhibition of ischaemic succinate accumulation and its oxidation after subsequent reperfusion is a potential therapeutic target to decrease ischaemia-reperfusion injury in a range of pathologies

Effects of calorie restriction on life span of microorganisms

Calorie restriction (CR) in microorganisms such as budding and fission yeasts has a robust and well-documented impact on longevity. In order to efficiently utilize the limited energy during CR, these organisms shift from primarily fermentative metabolism to mitochondrial respiration. Respiration activates certain conserved longevity factors such as sirtuins and is associated with widespread physiological changes that contribute to increased survival. However, the importance of respiration during CR-mediated longevity has remained controversial. The emergence of several novel metabolically distinct microbial models for longevity has enabled CR to be studied from new perspectives. The majority of CR and life span studies have been conducted in the primarily fermentative Crabtree-positive yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, but studies in primarily respiratory Crabtree-negative yeast and obligate aerobes can offer complementary insight into the more complex mammalian response to CR. Not only are microorganisms helping characterize a conserved cellular mechanism for CR-mediated longevity, but they can also directly impact mammalian metabolism as part of the natural gut flora. Here, we discuss the contributions of microorganisms to our knowledge of CR and longevity at the level of both the cell and the organism

Optimizing Dietary Restriction for Genetic Epistasis Analysis and Gene Discovery in C. elegans

Dietary restriction (DR) increases mammalian lifespan and decreases susceptibility to many age-related diseases. Lifespan extension due to DR is conserved across a wide range of species. Recent research has focused upon genetically tractable model organisms such as C. elegans to uncover the genetic mechanisms that regulate the response to DR, in the hope that this information will provide insight into the mammalian response and yield potential therapeutic targets. However, no consensus exists as to the best protocol to apply DR to C. elegans and potential key regulators of DR are protocol-specific. Here we define a DR method that better fulfills criteria required for an invertebrate DR protocol to mirror mammalian studies. The food intake that maximizes longevity varies for different genotypes and informative epistasis analysis with another intervention is only achievable at this ‘optimal DR’ level. Importantly therefore, the degree of restriction imposed using our method can easily be adjusted to determine the genotype-specific optimum DR level. We used this protocol to test two previously identified master regulators of DR in the worm. In contrast to previous reports, we find that DR can robustly extend the lifespan of worms lacking the AMP-activated protein kinase catalytic subunit AAK2 or the histone deacetylase SIR-2.1, highlighting the importance of first optimizing DR to identify universal regulators of DR mediated longevity

Tor1/Sch9-Regulated Carbon Source Substitution Is as Effective as Calorie Restriction in Life Span Extension

The effect of calorie restriction (CR) on life span extension, demonstrated in organisms ranging from yeast to mice, may involve the down-regulation of pathways, including Tor, Akt, and Ras. Here, we present data suggesting that yeast Tor1 and Sch9 (a homolog of the mammalian kinases Akt and S6K) is a central component of a network that controls a common set of genes implicated in a metabolic switch from the TCA cycle and respiration to glycolysis and glycerol biosynthesis. During chronological survival, mutants lacking SCH9 depleted extracellular ethanol and reduced stored lipids, but synthesized and released glycerol. Deletion of the glycerol biosynthesis genes GPD1, GPD2, or RHR2, among the most up-regulated in long-lived sch9Δ, tor1Δ, and ras2Δ mutants, was sufficient to reverse chronological life span extension in sch9Δ mutants, suggesting that glycerol production, in addition to the regulation of stress resistance systems, optimizes life span extension. Glycerol, unlike glucose or ethanol, did not adversely affect the life span extension induced by calorie restriction or starvation, suggesting that carbon source substitution may represent an alternative to calorie restriction as a strategy to delay aging

The physiological basis for genetic variation in water use efficiency and carbon isotope composition in Arabidopsis thaliana.

Ecologists and physiologists have documented extensive variation in water use efficiency (WUE) in Arabidopsis thaliana, as well as association of WUE with climatic variation. Here, we demonstrate correlations of whole-plant transpiration efficiency and carbon isotope composition (δ(13)C) among life history classes of A. thaliana. We also use a whole-plant cuvette to examine patterns of co-variation in component traits of WUE and δ(13)C. We find that stomatal conductance (g s) explains more variation in WUE than does A. Overall, there was a strong genetic correlation between A and g s, consistent with selection acting on the ratio of these traits. At a more detailed level, genetic variation in A was due to underlying variation in both maximal rate of carboxylation (V cmax) and maximum electron transport rate (Jmax). We also found strong effects of leaf anatomy, where lines with lower WUE had higher leaf water content (LWC) and specific leaf area (SLA), suggesting a role for mesophyll conductance (g m) in variation of WUE. We hypothesize that this is due to an effect through g m, and test this hypothesis using the abi4 mutant. We show that mutants of ABI4 have higher SLA, LWC, and g m than wild-type, consistent with variation in leaf anatomy causing variation in g m and δ(13)C. These functional data also add further support to the central, integrative role of ABI4 in simultaneously altering ABA sensitivity, sugar signaling, and CO2 assimilation. Together our results highlight the need for a more holistic approach in functional studies, both for more accurate annotation of gene function and to understand co-limitations to plant growth and productivity

Pleiotropy of FRIGIDA enhances the potential for multivariate adaptation.

An evolutionary response to selection requires genetic variation; however, even if it exists, then the genetic details of the variation can constrain adaptation. In the simplest case, unlinked loci and uncorrelated phenotypes respond directly to multivariate selection and permit unrestricted paths to adaptive peaks. By contrast, 'antagonistic' pleiotropic loci may constrain adaptation by affecting variation of many traits and limiting the direction of trait correlations to vectors that are not favoured by selection. However, certain pleiotropic configurations may improve the conditions for adaptive evolution. Here, we present evidence that the Arabidopsis thaliana gene FRI (FRIGIDA) exhibits 'adaptive' pleiotropy, producing trait correlations along an axis that results in two adaptive strategies. Derived, low expression FRI alleles confer a 'drought escape' strategy owing to fast growth, low water use efficiency and early flowering. By contrast, a dehydration avoidance strategy is conferred by the ancestral phenotype of late flowering, slow growth and efficient water use during photosynthesis. The dehydration avoidant phenotype was recovered when genotypes with null FRI alleles were transformed with functional alleles. Our findings indicate that the well-documented effects of FRI on phenology result from differences in physiology, not only a simple developmental switch