Engineering of itaconic acid pathway via co-localization of CadA and AcnA in recombinant Escherichia coli.

Itaconic acid is an excellent polymeric precursor with a wide range of industrial applications. The efficient production of itaconate from various renewable substrates was demonstrated by engineered Escherichia coli. However, limitation in the itaconic acid precursor supply was revealed by finding out the key intermediate of the tricarboxylic acid in the itaconic acid pathway. Efforts of enhancing the cis-aconitate flux and preserving the isocitrate pool to increase itaconic acid productivity are required. In this study, we introduce a synthetic protein scaffold system between CadA and AcnA to physically combine the two enzymes. Through the introduction of a synthetic protein scaffold, 2.1 g L-1 of itaconic acid was produced at pH 7 and 37 °C. By fermentation, 20.1 g L-1 for 48 h of itaconic acid was produced with a yield of 0.34 g g-1 glycerol. These results suggest that carbon flux was successfully increased itaconic acid productivity.

Investigation of CACNA1I Cav3.3 Dysfunction in Hemiplegic Migraine.

Familial hemiplegic migraine (FHM) is a severe neurogenetic disorder for which three causal genes, CACNA1A, SCN1A, and ATP1A2, have been implicated. However, more than 80% of referred diagnostic cases of hemiplegic migraine (HM) are negative for exonic mutations in these known FHM genes, suggesting the involvement of other genes. Using whole-exome sequencing data from 187 mutation-negative HM cases, we identified rare variants in the CACNA1I gene encoding the T-type calcium channel Cav3.3. Burden testing of CACNA1I variants showed a statistically significant increase in allelic burden in the HM case group compared to gnomAD (OR = 2.30, P = 0.00005) and the UK Biobank (OR = 2.32, P = 0.0004) databases. Dysfunction in T-type calcium channels, including Cav3.3, has been implicated in a range of neurological conditions, suggesting a potential role in HM. Using patch-clamp electrophysiology, we compared the biophysical properties of five Cav3.3 variants (p.R111G, p.M128L, p.D302G, p.R307H, and p.Q1158H) to wild-type (WT) channels expressed in HEK293T cells. We observed numerous functional alterations across the channels with Cav3.3-Q1158H showing the greatest differences compared to WT channels, including reduced current density, right-shifted voltage dependence of activation and inactivation, and slower current kinetics. Interestingly, we also found significant differences in the conductance properties exhibited by the Cav3.3-R307H and -Q1158H variants compared to WT channels under conditions of acidosis and alkalosis. In light of these data, we suggest that rare variants in CACNA1I may contribute to HM etiology.

Wheat transcriptomic responses to extended feeding by wheat curl mites.

The economic importance of wheat and its contribution to human and livestock diets has been already demonstrated. However, wheat production is impacted by pests that induce yield reductions. Among these pests, wheat curl mite (WCM, Aceria tosichella Keifer) impacts wheat all around the world. WCM are tiny pests that feed within the whorl of developing leaves, and their feeding causes leaf curling by preventing them from unfurling. The curling of the leaves provides a protective niche for the WCM. Additionally, WCM are also the vector of serious viruses in wheat. Little is known regarding the impact of the WCM on wheat transcriptome, and to date, only one article has been published describing the wheat transcriptomic changes after 1 day of WCM feeding. To better understand the wheat transcriptome variation after extended feeding by WCM [10 days post infestation (dpi)], we used an RNA-seq approach. We collected WCM-infested and uninfested leaves from two wheat cultivars: Byrd (WCM resistant) and Settler CL (WCM susceptible) at 10 dpi. Our transcriptomic analysis revealed the common and specific transcriptomic variations in WCM resistant and susceptible wheat cultivars, chromosome 3D specific location of the differentially expressed genes with functions involved in defense and stress response, and also identified the gene functions related to lipid signaling and membrane integrity, and phytohormone pathways potentially contributing to WCM resistance. Collectively, our study provides important insights on wheat defense mechanisms against WCM after extended feeding.

Chloroaluminate Ionic Liquid Immobilized on Magnetic Nanoparticles as a Heterogeneous Lewis Acidic Catalyst for the Friedel-Crafts Sulfonylation of Aromatic Compounds.

Chloroaluminate ionic liquid bound on magnetic nanoparticles (Fe3O4@O2Si[PrMIM]Cl·AlCl3) was prepared and used as a heterogenous Lewis acidic catalyst for the Friedel-Crafts sulfonylation of aromatic compounds with sulfonyl chlorides or p-toluenesulfonic anhydride. The catalyst's stability, efficiency, easy recovery, and high recyclability without considerable loss of catalytic capability after four recycles were evidence of its advantages. Furthermore, the stoichiometry, wide substrate scope, short reaction time, high yield of sulfones, and solvent-free reaction condition also made this procedure practical, ecofriendly, and economical.

Mimic microgravity effect on muscle transcriptome under ionizing radiation.

Spaceflight imposes the risk of skeletal muscle atrophy for astronauts. Two main factors of a spaceflight that results in deleterious effects are microgravity and cosmic rays in outer space. To study spaceflight-induced muscle atrophy with ground-based models, we performed two models of microgravity, tail suspension and denervation, in a low dose radiation environment and studied transcriptional changes in rat soleus muscle using microarrays. Soleus muscle from rats in the denervation group had greater expression changes compared to that found in rats from the tail suspension group. However, there was a very similar pattern of expression of differentially expressed genes (DEGs) in both models. In total, we identified 144 differentially expressed genes common in both models. Our study yielded two main findings. First, a large number of genes involved in energy metabolism were transcriptionally suppressed including those involved in fatty acid transport and beta-oxidation, and oxidative phosphorylation. Second, slow-twitch contractile protein encoding genes were down-regulated while there was an up-regulation in the fast-twitch type transcription. These results were consistent with other spaceflight studies on the effects on muscle cells, hence showed the potential of our ground-based models in studying spaceflight effects. The genes that might be involved in spaceflight effects will serve as candidate genes for future studies in understanding the mechanism of spaceflight-induced muscle atrophy and result in the development of effective countermeasures.

Efficient Itaconic acid production via protein-protein scaffold introduction between GltA, AcnA, and CadA in recombinant Escherichia coli.

Itaconic acid, which is a promising organic acid in synthetic polymers and some base-material production, has been produced by Aspergillus terreus fermentation at a high cost. The recombinant Escherichia coli that contained the cadA gene from A. terreus can produce itaconic acid but with low yield. By introducing the protein-protein scaffold between citrate synthesis, aconitase, and cis-aconitase decarboxylase, 5.7 g/L of itaconic acid was produced, which is 3.8-fold higher than that obtained with the strain without scaffold. The optimum pH and temperature for itaconic acid production were 8.5 and 30°C, respectively. When the competing metabolic network was inactivated by knock-out mutation, the itaconic acid concentration further increased, to 6.57 g/L.