Convergent changes in muscle metabolism depend on duration of high-altitude ancestry across Andean waterfowl

High-altitude environments require that animals meet the metabolic O2 demands for locomotion and thermogenesis in O2-thin air, but the degree to which convergent metabolic changes have arisen across independent high-altitude lineages or the speed at which such changes arise is unclear. We examined seven high-altitude waterfowl that have inhabited the Andes (3812–4806 m elevation) over varying evolutionary time scales, to elucidate changes in biochemical pathways of energy metabolism in flight muscle relative to low-altitude sister taxa. Convergent changes across high-altitude taxa included increased hydroxyacyl-coA dehydrogenase and succinate dehydrogenase activities, decreased lactate dehydrogenase, pyruvate kinase, creatine kinase, and cytochrome c oxidase activities, and increased myoglobin content. ATP synthase activity increased in only the longest established high-altitude taxa, whereas hexokinase activity increased in only newly established taxa. Therefore, changes in pathways of lipid oxidation, glycolysis, and mitochondrial oxidative phosphorylation are common strategies to cope with high-altitude hypoxia, but some changes require longer evolutionary time to arise.

Indications for a Central Role of Hexokinase Activity in Natural Variation of Heat Acclimation in Arabidopsis thaliana

Diurnal and seasonal changes of abiotic environmental factors shape plant performance and distribution. Changes of growth temperature and light intensity may vary significantly on a diurnal, but also on a weekly or seasonal scale. Hence, acclimation to a changing temperature and light regime is essential for plant survival and propagation. In the present study, we analyzed photosynthetic CO2 assimilation and metabolic regulation of the central carbohydrate metabolism in two natural accessions of Arabidopsis thaliana that originate from north western Russia and south Italy during exposure to heat and a combination of heat and high light. Our findings indicate that it is hardly possible to predict photosynthetic capacities under combined stress from single stress experiments. Further, capacities of hexose phosphorylation were found to be significantly lower in the Italian than in the Russian accession, which could explain an inverted sucrose-to-hexose ratio. Together with the finding of significantly stronger accumulation of anthocyanins under heat/high light, these observations indicate a central role of hexokinase activity in the stabilization of photosynthesis and carbohydrate metabolism during environmental changes.

Indications for a Central Role of Hexokinase Activity in Natural Variation of Heat Acclimation in Arabidopsis thaliana

Diurnal and seasonal changes of abiotic environmental factors shape plant performance and distribution. Changes of growth temperature and light intensity may vary significantly on a diurnal, but also on a weekly or seasonal scale. Hence, acclimation to a changing temperature and light regime is essential for plant survival and propagation. In the present study, we analyzed photosynthetic CO2 assimilation and metabolic regulation of the central carbohydrate metabolism in two natural accessions of Arabidopsis thaliana originating from Russia and south Italy during exposure to heat and a combination of heat and high light. Our findings indicate that it is hardly possible to predict photosynthetic capacities to fix CO2 under combined stress from single stress experiments. Further, capacities of hexose phosphorylation were found to be significantly lower in the Italian than in the Russian accession which could explain an inverted sucrose-to-hexose ratio. Together with the finding of significantly stronger accumulation of anthocyanins under heat/high light these observations indicate a central role of hexokinase activity in stabilization of photosynthetic capacities within a changing environment.

Cornifronone: A cadinane-type sesquiterpene from a mason bee (Osmia cornifrons)–derived Streptomyces sp

A new cadinane-type sesquiterpene named cornifronone is isolated from the body surface of a mason bee ( Osmia cornifrons)–derived Streptomyces sp. OC1611-8A. Its structure is identified by high-resolution electrospray ionization mass spectrometry data and nuclear magnetic resonance spectroscopic analysis. The absolute configuration of cornifronone was determined by electronic circular dichroism spectra calculations. Cornifronone inhibits hexokinase activity with a mean IC50 of 124.3 μM.

Polymerization in the actin ATPase clan regulates hexokinase activity in yeast

The actin fold is found in cytoskeletal polymers, chaperones, and various metabolic enzymes. Many actin-fold proteins, such as the carbohydrate kinases, do not polymerize. We found that Glk1, a Saccharomyces cerevisiae glucokinase, forms two-stranded filaments with ultrastructure that is distinct from that of cytoskeletal polymers. In cells, Glk1 polymerized upon sugar addition and depolymerized upon sugar withdrawal. Polymerization inhibits enzymatic activity; the Glk1 monomer-polymer equilibrium sets a maximum rate of glucose phosphorylation regardless of Glk1 concentration. A mutation that eliminated Glk1 polymerization alleviated concentration-dependent enzyme inhibition. Yeast containing nonpolymerizing Glk1 were less fit when growing on sugars and more likely to die when refed glucose. Glk1 polymerization arose independently from other actin-related filaments and may allow yeast to rapidly modulate glucokinase activity as nutrient availability changes.

Rapid radiochemical filter paper assay for determination of hexokinase activity and affinity for glucose-6-phosphate

Glucose phosphorylation by hexokinase (HK) is a rate-limiting step in glucose metabolism. Regulation of HK includes feedback inhibition by its product glucose-6-phosphate (G6P) and mitochondria binding. HK affinity for G6P is difficult to measure because its natural product (G6P) inhibits enzyme activity. HK phosphorylates several hexoses, and we have taken advantage of the fact that 2-deoxyglucose (2-DG)-6-phosphate does not inhibit HK activity. By this, we have developed a new method for rapid radiochemical analysis of HK activity with 2-DG as a substrate, which allows control of the concentrations of G6P to investigate HK affinity for inhibition by G6P. We verified that 2-DG serves as a substrate for the HK reaction with linear time and concentration dependency as well as expected maximal velocity and KM. This is the first simple assay that evaluates feedback inhibition of HK by its product G6P and provides a unique technique for future research evaluating the regulation of glucose phosphorylation under various physiological conditions. NEW & NOTEWORTHY Traditionally, hexokinase activity has been analyzed spectrophotometrically in which the product formation of glucose-6-phosphate (G6P) is analyzed by an indirect reaction coupled to NADPH formation during conversion of G6P to 6-P gluconolactone. By nature, this assay prevents measurements of hexokinase (HK) affinity for inhibition by G6P. We have developed a rapid radiochemical filter paper assay to study HK affinity for G6P by use of radiolabeled 2-deoxyglucose as substrate to study physiological regulation of HK affinity for G6P-induced inhibition.

Independent evolution of polymerization in the Actin ATPase clan regulates hexokinase activity

The actin protein fold is found in cytoskeletal polymers, chaperones, and various metabolic enzymes. Many actin-fold proteins, like the carbohydrate kinases, do not polymerize. We find that Glk1, aSaccharomyces cerevisiaeglucokinase, forms two-stranded filaments with unique ultrastructure, distinct from that of cytoskeletal polymers. In cells, Glk1 polymerizes upon sugar addition and depolymerizes upon sugar withdrawal. Glk1 polymerization inhibits its enzymatic activity, thus the Glk1 monomer-polymer equilibrium sets a maximum rate of glucose phosphorylation regardless of Glk1 concentration. A mutation eliminating Glk1 polymerization alleviates concentration-dependent enzyme inhibition, causing glucokinase activity to become unconstrained. Polymerization-based regulation of Glk1 activity serves an important functionin vivo: yeast containing non-polymerizing Glk1 are less fit when growing on sugars and more likely to die when refed glucose. Glucokinase polymerization arose within the ascomycete fungi and is conserved across a group of divergent (150-200 mya) yeast. We show that Glk1 polymerization arose independently from other actin-related filaments and allows yeast to rapidly modulate glucokinase activity as nutrient availability changes.One-sentence summaryYeast glucokinase activity is limited by its polymerization, which is critical for cell viability during glucose refeeding.