Comparative analysis of the zeta-crystallin/quinone reductase gene in guinea pig and mouse

zeta-Crystallin is a novel nicotinamide adenine dinucleotide phosphate:quinone reductase, present at enzymatic levels in various tissues of different species, which is highly expressed in the lens of some hystricomorph rodents and camelids. We report here the complementary DNA (cDNA) cloning of zeta-crystallin from liver libraries in guinea pig (Cavia porcellus), where zeta-crystallin is highly expressed in the lens, and in the laboratory mouse (Mus musculus), where expression in the lens occurs only at enzymatic levels. A 5' untranslated sequence different from the one previously reported for the guinea pig lens cDNA was found in these clones. We also report the isolation of genomic clones including the complete guinea pig zeta-crystallin gene and the 5' region of this gene in mouse. These results show the presence of two promoters in the guinea pig zeta-crystallin gene, one responsible for expression at enzymatic levels and the other responsible for the high expression in the lens. The guinea pig lens promoter is not present in the mouse gene. This is the first example in which the recruitment of an enzyme as a lens crystallin can be explained by the acquisition of an alternative lens- specific promoter.      

Calcium-dependent adenylate cyclase of pituitary tumor cells

Effects of Ca2+ and calmodulin on the adenylate cyclase activity of a prolactin and growth hormone-producing pituitary tumor cell strain (GH3) were examined. The adenylate cyclase activity of homogenates was stimulated approx. 60% by submicromolar free Ca2+ concentrations and inhibited by higher (microM range) concentrations of the cation. A 2-3-fold stimulation of the activity in response to Ca2+ was observed at physiologic concentrations of KCl, with both the stimulatory and inhibitory responses occurring at respectively higher free Ca2+ concentrations. Calmodulin in incubations at low KCl concentrations increased the enzyme activity at all Ca2+ concentrations tested. In incubations conducted at physiologic KCl concentrations, both the inhibitory and stimulatory responses to Ca2+ were shifted by calmodulin to lower respective concentrations of the cation, without significant change occurring in the maximal rate of enzymic activity at optimal free Ca2+ X Mg2+ concentrations in the incubation also influenced the Ca2+ concentration dependence of adenylate cyclase; at high Mg2+ more Ca2+ was required to obtain maximal activity. Trifluoperazine inhibited adenylate cyclase of GH3 cells only in the presence of Ca2+; as Ca2+ concentrations in the assay were increased, higher drug concentrations were required to inhibit the enzyme. Ca2+ was also observed to reduce the extent of enzyme destabilization which occurred during pretreatments at warm temperatures. Vasoactive intestinal polypeptide and phorbol myristate acetate, which stimulate prolactin secretion in intact GH3 cells, enhanced enzyme activity 4- and 2.5-fold, respectively, without added Ca2+. Increasing free Ca2+ concentrations reduced the enhancement by VIP and eliminated the stimulation by PMA.     

Metabolomic markers and physiological adaptations for high phosphate utilization efficiency in rice

Utilizing phosphate more efficiently is crucial for sustainable crop production. Highly efficient rice (Oryza sativa) cultivars have been identified and this study aims to identify metabolic markers associated with P utilization efficiency (PUE). P deficiency generally reduced leaf P concentrations and CO₂ assimilation rates but efficient cultivars were reducing leaf P concentrations further than inefficient ones while maintaining similar CO₂ assimilation rates. Adaptive changes in carbon metabolism were detected but equally in efficient and inefficient cultivar groups. Groups furthermore did not differ with respect to partial substitutions of phospholipids by sulfo- and galactolipids. Metabolites significantly more abundant in the efficient group, such as sinapate, benzoate and glucoronate, were related to antioxidant defence and may help alleviating oxidative stress caused by P deficiency. Sugar alcohols ribitol and threitol were another marker metabolite for higher phosphate efficiency as were several amino acids, especially threonine. Since these metabolites are not known to be associated with P deficiency, they may provide novel clues for the selection of more P efficient genotypes. In conclusion, metabolite signatures detected here were not related to phosphate metabolism but rather helped P efficient lines to keep vital processes functional under the adverse conditions of P starvation.     

Disruption of uridine homeostasis links liver pyrimidine metabolism to lipid accumulation

We report in this study an intrinsic link between pyrimidine metabolism and liver lipid accumulation utilizing a uridine phosphorylase 1 transgenic mouse model UPase1-TG. Hepatic microvesicular steatosis is induced by disruption of uridine homeostasis through transgenic overexpression of UPase1, an enzyme of the pyrimidine catabolism and salvage pathway. Microvesicular steatosis is also induced by the inhibition of dihydroorotate dehydrogenase (DHODH), an enzyme of the de novo pyrimidine biosynthesis pathway. Interestingly, uridine supplementation completely suppresses microvesicular steatosis in both scenarios. The effective concentration (EC₅₀) for uridine to suppress microvesicular steatosis is approximately 20 µM in primary hepatocytes of UPase1-TG mice. We find that uridine does not have any effect on in vitro DHODH enzymatic activity. On the other hand, uridine supplementation alters the liver NAD⁺/NADH and NADP⁺/NADPH ratios and the acetylation profile of metabolic, oxidation-reduction, and antioxidation enzymes. Protein acetylation is emerging as a key regulatory mechanism for cellular metabolism. Therefore, we propose that uridine suppresses fatty liver by modulating the liver protein acetylation profile. Our findings reveal a novel link between uridine homeostasis, pyrimidine metabolism, and liver lipid metabolism.     

Low-temperature tolerance of the Antarctic species Deschampsia antarctica: A complex metabolic response associated with nutrient remobilization

The species Deschampsia antarctica (DA) is one of the only two native vascular species that live in Antarctica. We performed ecophysiological, biochemical, and metabolomic studies to investigate the responses of DA to low temperature. In parallel, we assessed the responses in a non-Antarctic reference species (Triticum aestivum [TA]) from the same family (Poaceae). At low temperature (4°C), both species showed lower photosynthetic rates (reductions were 70% and 80% for DA and TA, respectively) and symptoms of oxidative stress but opposite responses of antioxidant enzymes (peroxidases and catalase). We employed fused least absolute shrinkage and selection operator statistical modelling to associate the species-dependent physiological and antioxidant responses to primary metabolism. Model results for DA indicated associations with osmoprotection, cell wall remodelling, membrane stabilization, and antioxidant secondary metabolism (synthesis of flavonols and phenylpropanoids), coordinated with nutrient mobilization from source to sink tissues (confirmed by elemental analysis), which were not observed in TA. The metabolic behaviour of DA, with significant changes in particular metabolites, was compared with a newly compiled multispecies dataset showing a general accumulation of metabolites in response to low temperatures. Altogether, the responses displayed by DA suggest a compromise between catabolism and maintenance of leaf functionality.     

Ovarian activity and plasma sex steroid levels of Distichodus antonii in relation to environmental conditions in the upper basin of the Congo River

Distichodus antonii is an endemic fish species of the Congo River basin in which the stocks of wild populations are threatened by overfishing pressure. Knowledge of its reproductive biology would be useful in consideration of conservation and management options for the species. Therefore, this study investigated changes in ovarian activity and levels of steroid profiles in wild populations in relation to variation in temperature and rainfall. Adult females (n = 101, body weight of 3 183 ± 14.75 g, SE) were captured monthly over one year (2013–2014). Apart from evaluation of oocyte diameters and gonad developmental stages, gonado-, hepato-, lipososomatic indices (GSI, HSI, LSI) and plasma levels of sex steroids (testosterone-T, estradiol-17β-E2) were determined. The results suggested a synchronous development of oocytes with two annual reproductive seasons over the one-year study. Plasma T and E2 levels peaked during spawning periods likely reflecting active oogenesis. The highest values of morphosomatic indices were observed during the longest rainfall period in September, and were associated with high steroidogenic activity evidenced by increased E2 production. In addition, more vitellogenic oocytes (September and October) were observed during the latter season than during the short rainy season (in May).     

The LysM Receptor-Like Kinase LysM RLK1 Is Required to Activate Defense and Abiotic-Stress Responses Induced by Overexpression of Fungal Chitinases in Arabidopsis Plants

Application of crab shell chitin or pentamer chitin oligosaccharide to Arabidopsis seedlings increased tolerance to salinity in wild-type but not in knockout mutants of the LysM Receptor-Like Kinase1 (CERK1/LysM RLK1) gene, known to play a critical role in signaling defense responses induced by exogenous chitin. Arabidopsis plants overexpressing the endochitinase chit36 and hexoaminidase excy1 genes from the fungus Trichoderma asperelleoides T203 showed increased tolerance to salinity, heavy-metal stresses, and Botrytis cinerea infection. Resistant lines, overexpressing fungal chitinases at different levels, were outcrossed to lysm rlk1 mutants. Independent homozygous hybrids lost resistance to biotic and abiotic stresses, despite enhanced chitinase activity. Expression analysis of 270 stress-related genes, including those induced by reactive oxygen species (ROS) and chitin, revealed constant up-regulation (at least twofold) of 10 genes in the chitinase-overexpressing line and an additional 76 salt-induced genes whose expression was not elevated in the lysm rlk1 knockout mutant or the hybrids harboring the mutation. These findings elucidate that chitin-induced signaling mediated by LysM RLK1 receptor is not limited to biotic stress response but also encompasses abiotic-stress signaling and can be conveyed by ectopic expression of chitinases in plants.