Gluconeogenesis in Plants: A Key Interface between Organic Acid/Amino Acid/Lipid and Sugar Metabolism

Gluconeogenesis is a key interface between organic acid/amino acid/lipid and sugar metabolism. The aims of this article are four-fold. First, to provide a concise overview of plant gluconeogenesis. Second, to emphasise the widespread occurrence of gluconeogenesis and its utilisation in diverse processes. Third, to stress the importance of the vacuolar storage and release of Krebs cycle acids/nitrogenous compounds, and of the role of gluconeogenesis and malic enzyme in this process. Fourth, to outline the contribution of fine control of enzyme activity to the coordinate-regulation of gluconeogenesis and malate metabolism, and the importance of cytosolic pH in this.

Quantitative Proteomics and Relative Enzymatic Activities Reveal Different Mechanisms in Two Peanut Cultivars (Arachis hypogaea L.) Under Waterlogging Conditions

Peanut is an important oil and economic crop in China. The rainy season (April–June) in the downstream Yangtze River in China always leads to waterlogging, which seriously affects plant growth and development. Therefore, understanding the metabolic mechanisms under waterlogging stress is important for future waterlogging tolerance breeding in peanut. In this study, waterlogging treatment was carried out in two different peanut cultivars [Zhonghua 4 (ZH4) and Xianghua08 (XH08)] with different waterlogging tolerance. The data-independent acquisition (DIA) technique was used to quantitatively identify the differentially accumulated proteins (DAPs) between two different cultivars. Meanwhile, the functions of DAPs were predicted, and the interactions between the hub DAPs were analyzed. As a result, a total of 6,441 DAPs were identified in ZH4 and its control, of which 49 and 88 DAPs were upregulated and downregulated under waterlogging stress, respectively, while in XH08, a total of 6,285 DAPs were identified, including 123 upregulated and 114 downregulated proteins, respectively. The hub DAPs unique to the waterlogging-tolerant cultivar XH08 were related to malate metabolism and synthesis, and the utilization of the glyoxylic acid cycle, such as L-lactate dehydrogenase, NAD+-dependent malic enzyme, aspartate aminotransferase, and glutamate dehydrogenase. In agreement with the DIA results, the alcohol dehydrogenase and malate dehydrogenase activities in XH08 were more active than ZH4 under waterlogging stress, and lactate dehydrogenase activity in XH08 was prolonged, suggesting that XH08 could better tolerate waterlogging stress by using various carbon sources to obtain energy, such as enhancing the activity of anaerobic respiration enzymes, catalyzing malate metabolism and the glyoxylic acid cycle, and thus alleviating the accumulation of toxic substances. This study provides insight into the mechanisms in response to waterlogging stress in peanuts and lays a foundation for future molecular breeding targeting in the improvement of peanut waterlogging tolerance, especially in rainy area, and will enhance the sustainable development in the entire peanut industry.

Engineering cooperation in an anaerobic co-culture

Over the past century, microbiologists have studied organisms in pure culture, yet it is becoming increasingly apparent that the majority of biological processes rely on multispecies cooperation and interaction. While little is known about how such interactions permit cooperation, even less is known about how cooperation arises. To study the emergence of cooperation in the laboratory, we constructed both a commensal community and an obligate mutualism using the previously non-interacting bacteria Shewanella oneidensis and Geobacter sulfurreducens. Incorporation of a glycerol utilization plasmid (pGUT2) enabled S. oneidensis to metabolize glycerol and produce acetate as a carbon source for G. sulfurreducens establishing a cross-feeding, commensal co-culture. In the commensal co-culture, both species coupled oxidative metabolism to the respiration of fumarate as the terminal electron acceptor. Deletion of the gene encoding fumarate reductase in the S. oneidensis pGUT2 strain shifted the co-culture with G. sulfurreducens to an obligate mutualism where neither species could grow in absence of the other. A shift in metabolic strategy from glycerol catabolism to malate metabolism was associated with obligate co-culture growth. Further targeted deletions in malate uptake and acetate generation pathways in S. oneidensis significantly inhibited co-culture growth with G. sulfurreducens. The engineered co-culture between S. oneidensis and G. sulfurreducens provides a model laboratory system to study the emergence of cooperation in bacterial communities, and the shift in metabolic strategy observed in the obligate co-culture highlights the importance of genetic change in shaping microbial interactions in the environment. Importance Microbes seldom live alone in the environment, yet this scenario is approximated in the vast majority of pure-culture laboratory experiments. Here we develop an anaerobic co-culture system to begin understanding microbial physiology in a more complex setting, but also to determine how anaerobic microbial communities can form. Using synthetic biology, we generated a co-culture system where the facultative anaerobe Shewanella oneidensis consumes glycerol and provides acetate to the strict anaerobe Geobacter sulfurreducens. In the commensal system, growth of G. sulfurreducens is dependent on the presence of S. oneidensis. To generate an obligate co-culture, where each organism requires the other, we eliminated the ability of S. oneidensis to respire fumarate. An unexpected shift in metabolic strategy from glycerol catabolism to malate metabolism was observed in the obligate co-culture. Our work highlights how metabolic landscapes can be expanded in multi-species communities and provides a system to evaluate the evolution of cooperation under anaerobic conditions.

The localization of nitrite reductase, glutamate synthase and malate metabolism enzymes in Pisum arvense L. roots

Centrifugation of a homogenate made from <em>Pisum arvense</em> L. roots in a sucrose density gradient enabled the separation of the plastid fraction from mitochondria and microsomes. The presence of nitrite reductase and glutamate synthase was demonstrated in the plastids. Malic enzyme activity was not linked with any organelle fraction and was found only in the cytosol. High malate dehydrogenase activity was found in the mitochondria fraction, although its activity was also determined in plastids. The results suggest that malic acid metabolism in plastids may be the source of reduced pyridine nucleotides for reactions catalysed by nitrite reductase and glutamate synthase.

Malic Enzyme and Malolactic Enzyme Pathways Are Functionally Linked but Independently Regulated in Lactobacillus casei BL23

ABSTRACTLactobacillus caseiis the only lactic acid bacterium in which two pathways forl-malate degradation have been described: the malolactic enzyme (MLE) and the malic enzyme (ME) pathways. Whereas the ME pathway enablesL. caseito grow onl-malate, MLE does not support growth. Themlegene cluster consists of three genes encoding MLE (mleS), the putativel-malate transporter MleT, and the putative regulator MleR. Themaegene cluster consists of four genes encoding ME (maeE), the putative transporter MaeP, and the two-component system MaeKR. Since both pathways compete for the same substrate, we sought to determine whether they are coordinately regulated and their role inl-malate utilization as a carbon source. Transcriptional analyses revealed that themleandmaegenes are independently regulated and showed that MleR acts as an activator and requires internalization ofl-malate to induce the expression ofmlegenes. Notwithstanding, bothl-malate transporters were required for maximall-malate uptake, although only anmleTmutation caused a growth defect onl-malate, indicating its crucial role inl-malate metabolism. However, inactivation of MLE resulted in higher growth rates and higher final optical densities onl-malate. The limited growth onl-malate of the wild-type strain was correlated to a rapid degradation of the availablel-malate tol-lactate, which cannot be further metabolized. Taken together, our results indicate thatL. caseil-malate metabolism is not optimized for utilization ofl-malate as a carbon source but for deacidification of the medium by conversion ofl-malate intol-lactate via MLE.