Stepwise reduction of the culture redox potential allows the analysis of microaerobic metabolism and photosynthetic membrane synthesis in Rhodospirillum rubrum

Bacterial growth under oxygen‐limited (microaerobic) conditions is often accompanied by phenomena of great interest for fundamental research and industrial application. The microaerobic lifestyle of anoxygenic photosynthetic bacteria like Rhodospirillum rubrum harbors such a phenomenon, as it allows the formation of photosynthetic membranes and related interesting products without light. However, due to the technical difficulties in process control of microaerobic cultivations and the limited sensitivity of available oxygen sensors, the analysis of microaerobic growth and physiology is still underrepresented in current research. The main focus of the present study was to establish an experimental set‐up for the systematic study of physiological processes, associated with the growth of R. rubrum under microaerobic conditions in the dark. For this purpose, we introduce a robust and reliable microaerobic process control strategy, which applies the culture redox potential (CRP) for assessing different degrees of oxygen limitation in bioreactor cultivations. To describe the microaerobic growth behavior of R. rubrum cultures for each of these defined CRP reduction steps, basic growth parameters were experimentally determined. Flux variability analysis provided an insight into the metabolic activity of the TCA cycle and implied its connection to the respiratory capacity of the cells. In this context, our results suggest that microaerobic growth of R. rubrum can be described as an oxygen‐activated cooperative mechanism. The present study thus contributes to the investigation of metabolic and regulatory events responsible for the redox‐sensitive formation of photosynthetic membranes in facultative photosynthetic bacteria. Furthermore, the introduced microaerobic cultivation setup should be generally applicable for any microbial system of interest which can be cultivated in common stirred‐tank bioreactors. Biotechnol. Bioeng. 2013; 110: 573–585. © 2012 Wiley Periodicals, Inc.     

Spatial memory deficits in maternal iron deficiency paradigms are associated with altered glucocorticoid levels

"The goal of this study was to examine the effect of maternal iron deficiency on the developing hippocampus in order to define a developmental window for this effect, and to see whether iron deficiency causes changes in glucocorticoid levels. The study was carried out using pre-natal, post-natal, and pre + post-natal iron deficiency paradigm. Iron deficient pregnant dams and their pups displayed elevated corticosterone which, in turn, differentially affected glucocorticoid receptor (GR) expression in the CA1 and the dentate gyrus. Brain Derived Neurotrophic Factor (BDNF) was reduced in the hippocampi of pups following elevated corticosterone levels. Reduced neurogenesis at P7 was seen in pups born to iron deficient mothers, and these pups had reduced numbers of hippocampal pyramidal and granule cells as adults. Hippocampal subdivision volumes also were altered. The structural and molecular defects in the pups were correlated with radial arm maze performance; reference memory function was especially affected. Pups from dams that were iron deficient throughout pregnancy and lactation displayed the complete spectrum of defects, while pups from dams that were iron deficient only during pregnancy or during lactation displayed subsets of defects. These findings show that maternal iron deficiency is associated with altered levels of corticosterone and GR expression, and with spatial memory deficits in their pups."     

我国亚热带毛竹林CO2通量的变异特征

2010年12月至2011年11月,利用涡度相关技术研究了我国亚热带(浙江)毛竹林生态系统的CO₂通量,分析了毛竹林净生态系统交换量(NEE)、生态系统呼吸量(RE)和生态系统总交换量(GEE)的变化.结果表明: 研究期间,毛竹林各月的NEE均为负值,7月最大,为-99.33 g C·m^-2,11月最小,仅-23.49 g C·m^-2,其变化曲线呈双峰型.各月CO₂通量平均日变化差异明显,9月最大,为-0.60 g CO₂·m^-2·s^-1,1月最小,为-0.30 g CO₂·m^-2·s^-1,且在NEE正负转换的时间点上呈明显的季节变化特征;全年RE呈单峰型变化,夏季最高、冬季最低,夜间RE与土壤温度呈极显著正相关.全年NEE、RE和GEE分别为-668.40、932.55和-1600.95 g C·m^-2·a^-1,NEE占GEE的41.8%.与其他生态系统相比,毛竹林的固碳能力极强.     

A dynamic metabolic flux balance based model of fed‐batch fermentation of bordetella pertussis

A mathematical model based on a dynamic metabolic flux balance (DMFB) is developed for a process of fed‐batch fermentation of Bordetella pertussis. The model is based on the maximization of growth rate at each time interval subject to stoichiometric constraints. The model is calibrated and verified with experimental data obtained in two different bioreactor experimental systems. It was found that the model calibration was mostly sensitive to the consumption or production rates of tyrosine and, for high supplementation rates, to the consumption rate of glutamate. Following this calibration the model correctly predicts biomass and by‐products concentrations for different supplementation rates. Comparisons of model predictions to oxygen uptake and carbon emission rates measurements indicate that the TCA cycle is fully functional. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 520–531, 2013     

Decreasing the Roughage: Concentrate Ratio of a Diet to Determine the Critical Roughage Part for Beef Cattle

The critical roughage part (CRP) of 2 diet types was determined in a cross-over design with 6 double-muscled and 6 normally conformated Belgian Blue bulls fitted with rumen cannulae. The roughage:concentrate ratio was lowered weekly until signs of a lack of physical structure were observed. For diet 1, consisting of maize silage and concentrates, the initial proportion of maize silage was 25% of DM but it decreased weekly with 5% units of DM. For the second diet, consisting of wheat straw and concentrate, 12% straw (DM basis) was provided during the first week and thereafter the proportion of straw decreased weekly with 3% units of DM. Several directly observable parameters (rumen pH, feed intake, bloat, faecal consistency) were evaluated weekly for each bull. Apart from these direct indicators of acidosis, also other parameters, whose results were only available after the end of the trial, were determined (volatile fatty acid profile, lactic acid concentration, chewing time). The roughage part between the part fed when signs of a lack of physical structure was first observed and the part that was fed the week before, was considered as the CRP. Most animals showed no acute signs of clinical acidosis (directly observable parameters) and finished the trial on a 100% concentrate diet. However, in sacco rumen DM-degradabilities of maize silage, grass silage and wheat grain was depressed considerably when low roughage diets were fed. Based on all observed parameters, the mean CRP was calculated to be 14.7% for diet 1 and 8.1% for diet 2. The beef type (double-muscled or not) had no influence on the CRP.     

Protein design in systems metabolic engineering for industrial strain development

Accelerating the process of industrial bacterial host strain development, aimed at increasing productivity, generating new bio-products or utilizing alternative feedstocks, requires the integration of complementary approaches to manipulate cellular metabolism and regulatory networks. Systems metabolic engineering extends the concept of classical metabolic engineering to the systems level by incorporating the techniques used in systems biology and synthetic biology, and offers a framework for the development of the next generation of industrial strains. As one of the most useful tools of systems metabolic engineering, protein design allows us to design and optimize cellular metabolism at a molecular level. Here, we review the current strategies of protein design for engineering cellular synthetic pathways, metabolic control systems and signaling pathways, and highlight the challenges of this subfield within the context of systems metabolic engineering.     

Wild-type rabies virus induces autophagy in human and mouse neuroblastoma cell lines

Different rabies virus (RABV) strains have their own biological characteristics, but little is known about their respective impact on autophagy. Therefore, we evaluated whether attenuated RABV HEP-Flury and wild-type RABV GD-SH-01 strains triggered autophagy. We found that GD-SH-01 infection significantly increased the number of autophagy-like vesicles, the accumulation of enhanced green fluorescent protein (EGFP)-LC3 fluorescence puncta and the conversion of LC3-I to LC3-II, while HEP-Flury was not able to induce this phenomenon. When evaluating autophagic flux, we found that GD-SH-01 infection triggers a complete autophagic response in the human neuroblastoma cell line (SK), while autophagosome fusion with lysosomes was inhibited in a mouse neuroblastoma cell line (NA). In these cells, GD-SH-01 led to apoptosis and mitochondrial dysfunction while triggering autophagy, and apoptosis could be decreased by enhancing autophagy. To further identify the virus constituent causing autophagy, 5 chimeric recombinant viruses carrying single genes of HEP-Flury instead of those of GD-SH-01 were rescued. While the HEP-Flury virus carrying the wild-type matrix protein (M) gene of RABV triggered LC3-I to LC3-II conversion in SK and NA cells, replacement of genes of nucleoprotein (N), phosphoprotein (P) and glycoprotein (G) produced only minor autophagy. But no one single structural protein of GD-SH-01 induced autophagy. Moreover, the AMPK signaling pathway was activated by GD-SH-01 in SK. Therefore, our data provide strong evidence that autophagy is induced by GD-SH-01 and can decrease apoptosis in vitro. Furthermore, the M gene of GD-SH-01 may cooperatively induce autophagy.     

Global analysis of cellular protein flux quantifies the selectivity of basal autophagy

In eukaryotic cells, the macroautophagy pathway has been implicated in the degradation of long-lived proteins and damaged organelles. Although it has been demonstrated that macroautophagy can selectively degrade specific targets, its contribution to the basal turnover of cellular proteins had previously not been quantified on proteome-wide scales. In a recent study, we utilized dynamic proteomics to provide a global comparison of protein half-lives between wild-type and autophagy-deficient cells. Our results indicated that in quiescent fibroblasts, macroautophagy contributes to the basal turnover of a substantial fraction of the proteome. However, the contribution of macroautophagy to constitutive protein turnover is variable within the proteome. The methodology outlined in the study provides a global strategy for quantifying the selectivity of basal macroautophagy.