Genome-scale modeling for Bacillus coagulans to understand the metabolic characteristics

Lactic acid is widely used in many industries, especially in the production of poly-lactic acid. Bacillus coagulans is a promising lactic acid producer in industrial fermentation due to its thermophilic property. In this study, we developed the first genome-scale metabolic model (GEM) of B. coagulans iBag597, together with an enzyme-constrained model ec-iBag597. We measured strain-specific biomass composition and integrated the data into a biomass equation. Then, we validated iBag597 against experimental data generated in this study, including amino acid requirements and carbon source utilization, showing that simulations were generally consistent with the experimental results. Subsequently, we carried out chemostats to investigate the effects of specific growth rate and culture pH on metabolism of B. coagulans. Meanwhile, we used iBag597 to estimate the intracellular metabolic fluxes for those conditions. The results showed that B. coagulans was capable of generating ATP via multiple pathways, and switched among them in response to various conditions. With ec-iBag597, we estimated the protein cost and protein efficiency for each ATP-producing pathway to investigate the switches. Our models pave the way for systems biology of B. coagulans, and our findings suggest that maintaining a proper growth rate and selecting an optimal pH are beneficial for lactate fermentation.     

A predominantly nuclear protein affecting cytoplasmic localization of beta-actin mRNA in fibroblasts and neurons.

The localization of beta-actin mRNA to the leading lamellae of chicken fibroblasts and neurite growth cones of developing neurons requires a 54-nt localization signal (the zipcode) within the 3' untranslated region. In this study we have identified and isolated five proteins binding to the zipcode. One of these we previously identified as zipcode binding protein (ZBP)1, a 4-KH domain protein. A second is now investigated in detail: a 92-kD protein, ZBP2, that is especially abundant in extracts from embryonic brain. We show that ZBP2 is a homologue of the human hnRNP protein, KSRP, that appears to mediate pre-mRNA splicing. However, ZBP2 has a 47-amino acid (aa) sequence not present in KSRP. Various portions of ZBP2 fused to GFP indicate that the protein most likely shuttles between the nucleus and the cytoplasm, and that the 47-aa insert promotes the nuclear localization. Expression of a truncated ZBP2 inhibits the localization of beta-actin mRNA in both fibroblast and neurons. These data suggest that ZBP2, although predominantly a nuclear protein, has a role in the cytoplasmic localization of beta-actin mRNA.     

Activation of Toll-like Receptor 4 (TLR4) Attenuates Adaptive Thermogenesis via Endoplasmic Reticulum Stress

Adaptive thermogenesis is the cellular process transforming chemical energy into heat in response to cold. A decrease in adaptive thermogenesis is a contributing factor to obesity. However, the molecular mechanisms responsible for the compromised adaptive thermogenesis in obese subjects have not yet been elucidated. In this study we hypothesized that Toll-like receptor 4 (TLR4) activation and subsequent inflammatory responses are key regulators to suppress adaptive thermogenesis. To test this hypothesis, C57BL/6 mice were either fed a palmitate-enriched high fat diet or administered with chronic low-dose LPS before cold acclimation. TLR4 stimulation by a high fat diet or LPS were both associated with reduced core body temperature and heat release. Impairment of thermogenic activation was correlated with diminished expression of brown-specific markers and mitochondrial dysfunction in subcutaneous white adipose tissue (sWAT). Defective sWAT browning was concomitant with elevated levels of endoplasmic reticulum (ER) stress and autophagy. Consistently, TLR4 activation by LPS abolished cAMP-induced up-regulation of uncoupling protein 1 (UCP1) in primary human adipocytes, which was reversed by silencing of C/EBP homologous protein (CHOP). Moreover, the inactivation of ER stress by genetic deletion of CHOP or chemical chaperone conferred a resistance to the LPS-induced suppression of adaptive thermogenesis. Collectively, our data indicate the existence of a novel signaling network that links TLR4 activation, ER stress, and mitochondrial dysfunction, thereby antagonizing thermogenic activation of sWAT. Our results also suggest that TLR4/ER stress axis activation may be a responsible mechanism for obesity-mediated defective brown adipose tissue activation.     

Aldosterone induction of hepatic stellate cell contraction through activation of RhoA/ROCK-2 signaling pathway

The RhoA/ROCK-2 signaling pathway is necessary for activated hepatic stellate cell (HSC) contraction. HSC contraction plays an important role in the pathogenesis of cirrhosis and portal hypertension. This study investigated whether aldosterone contributes to HSC contraction by activation of the RhoA/ROCK-2 signaling pathway. Primary HSCs were isolated from Sprague-Dawley rats via in situ pronase/collagenase perfusion. We found that aldosterone enhanced the contraction of a collagen lattice seeded with HSCs. This induced contraction was suppressed by the mineralcorticoid receptor (MR) inhibitor spironolactone, the ROCK-2 inhibitor Y27632, and the angiotensin II type 1 receptor (AT(1)R) inhibitor irbesartan. Moreover, actin fiber staining showed that aldosterone significantly increased actin fiber formation in HSCs. Pre-incubating with spironolactone, Y27632, or irbesartan inhibited the aldosterone-induced actin fiber reorganization. Molecularly, the effect of aldosterone on activation of HSC contraction was mediated by phosphorylated myosin light chain (P-MLC) through the RhoA/ROCK-2 signaling pathway. All these inhibitors had the ability to block aldosterone-induced protein expressions in the RhoA/ROCK-2/P-MLC cascade in HSCs. Taken together, our current study suggests that aldosterone induces contraction of activated HSCs through the activation of the RhoA/ROCK-2 signaling pathway. This finding may provide a potential therapeutic target for control of cirrhosis and portal hypertension.     

The first complete plastome sequence of the basal asterid family Styracaceae (Ericales) reveals a large inversion

Plastome sequences are rich sources of information for resolving difficult phylogenetic relationships and provide genomic data for conservation studies. Here, the complete plastome sequence of Alniphyllum eberhardtii Guillaumin is reported, representing the first plastome of the basal asterid family Styracaceae (Ericales). The plastome is 155,384 bp in length and contains 79 protein-coding genes, 30 tRNA genes and 4 rRNA genes, totaling 113 unique genes with 19 genes in the inverted repeat region. Unusual features of the plastome include the presence a large 20-kb inversion in the Large Single-Copy region, the pseudogenization of the accD gene, and the loss of the second intron from clpP. The 20-kb inversion includes 14 genes and has not been previously reported in other Ericales plastomes. Thirty-nine plastid simple sequence repeats (SSRs) that may provide genetic resources for the conservation of this economically import timber plant are characterized. Phylogenetic results inferred from ML and MP analyses of 66 plastid genes and 26 taxa reveal that the Styracaceae are sister to a clade including Actinidiaceae and Ericaceae and suggest that complete plastomes are likely to be very helpful in resolving the basal relationships among Ericales families, which have resisted resolution in smaller phylogenetic data sets.     

Formation of soil organic carbon pool is regulated by the structure of dissolved organic matter and microbial carbon pump efficacy: A decadal study comparing different carbon management strategies

To achieve long-term increases in soil organic carbon (SOC) storage, it is essential to understand the effects of carbon management strategies on SOC formation pathways, particularly through changes in microbial necromass carbon (MNC) and dissolved organic carbon (DOC). Using a 14-year field study, we demonstrate that both biochar and maize straw lifted the SOC ceiling, but through different pathways. Biochar, while raising SOC and DOC content, decreased substrate degradability by increasing carbon aromaticity. This resulted in suppressed microbial abundance and enzyme activity, which lowered soil respiration, weakened in vivo turnover and ex vivo modification for MNC production (i.e., low microbial carbon pump “efficacy”), and led to lower efficiency in decomposing MNC, ultimately resulting in the net accumulation of SOC and MNC. In contrast, straw incorporation increased the content and decreased the aromaticity of SOC and DOC. The enhanced SOC degradability and soil nutrient content, such as total nitrogen and total phosphorous, stimulated the microbial population and activity, thereby boosting soil respiration and enhancing microbial carbon pump “efficacy” for MNC production. The total C added to biochar and straw plots were estimated as 27.3–54.5 and 41.4 Mg C ha⁻¹, respectively. Our results demonstrated that biochar was more efficient in lifting the SOC stock via exogenous stable carbon input and MNC stabilization, although the latter showed low “efficacy”. Meanwhile, straw incorporation significantly promoted net MNC accumulation but also stimulated SOC mineralization, resulting in a smaller increase in SOC content (by 50%) compared to biochar (by 53%–102%). The results address the decadal-scale effects of biochar and straw application on the formation of the stable organic carbon pool in soil, and understanding the causal mechanisms can allow field practices to maximize SOC content.     

Assessment of selenium bioavailability from naturally produced high-selenium soy foods in selenium-deficient rats

We assessed the bioavailability of selenium (Se) from a protein isolate and tofu (bean curd) prepared from naturally produced high-Se soybeans. The Se concentrations of the soybeans, the protein isolate and tofu were 5.2 ± 0.2, 11.4 ± 0.1 and 7.4 ± 0.1 mg/kg, respectively. Male weanling Sprague–Dawley rats were depleted of Se by feeding them a 30% Torula yeast-based diet (4.1 μg Se/kg) for 56 days, and then they were replenished with Se for an additional 50 days by feeding them the same diet containing 14, 24 or 30 μg Se/kg from the protein isolate or 13, 23 or 31 μg Se/kg from tofu, respectively. l-Selenomethionine (SeMet) was used as a reference. Selenium bioavailability was determined on the basis of the restoration of Se-dependent enzyme activities and tissue Se concentrations in Se-depleted rats, comparing those responses for the protein isolate and tofu to those for SeMet by using a slope-ratio method. Dietary supplementation with the protein isolate or tofu resulted in linear or log-linear, dose-dependent increases in glutathione peroxidase activities in blood and liver and in thioredoxin reductase activity in liver. Furthermore, supplementation with the protein isolate or tofu resulted in linear or log-linear, dose-dependent increases in the Se concentrations of plasma, liver, muscle and kidneys. These results indicated an overall bioavailability of approximately 101% for Se from the protein isolate and 94% from tofu, relative to SeMet. We conclude that Se from naturally produced high-Se soybeans is highly bioavailable in this model and that high-Se soybeans may be a good dietary source of Se.     

CD1 molecules efficiently present antigen in immature dendritic cells and traffic independently of MHC class II during dendritic cell maturation

Upon exposure to Ag and inflammatory stimuli, dendritic cells (DCs) undergo a series of dynamic cellular events, referred to as DC maturation, that involve facilitated peptide Ag loading onto MHC class II molecules and their subsequent transport to the cell surface. Besides MHC molecules, human DCs prominently express molecules of the CD1 family (CD1a, -b, -c, and -d) and mediate CD1-dependent presentation of lipid and glycolipid Ags to T cells, but the impact of DC maturation upon CD1 trafficking and Ag presentation is unknown. Using monocyte-derived immature DCs and those stimulated with TNF-alpha for maturation, we observed that none of the CD1 isoforms underwent changes in intracellular trafficking that mimicked MHC class II molecules during DC maturation. In contrast to the striking increase in surface expression of MHC class II on mature DCs, the surface expression of CD1 molecules was either increased only slightly (for CD1b and CD1c) or decreased (for CD1a). In addition, unlike MHC class II, DC maturation-associated transport from lysosomes to the plasma membrane was not readily detected for CD1b despite the fact that both molecules were prominently expressed in the same MIIC lysosomal compartments before maturation. Consistent with this, DCs efficiently presented CD1b-restricted lipid Ags to specific T cells similarly in immature and mature DCs. Thus, DC maturation-independent pathways for lipid Ag presentation by CD1 may play a crucial role in host defense, even before DCs are able to induce maximum activation of peptide Ag-specific T cells.