Studies on substrate utilisation in l-valine-producing Corynebacterium glutamicum strains deficient in pyruvate dehydrogenase complex

The pyruvate dehydrogenase complex was deleted to increase precursor availability in Corynebacterium glutamicum strains overproducing l-valine. The resulting auxotrophy is treated by adding acetate in addition glucose for growth, resulting in the puzzling fact of gluconeogenic growth with strongly reduced glucose uptake in the presence of acetate in the medium. This result was proven by intracellular metabolite analysis and labelling experiments. To increase productivity, the SugR protein involved in negative regulation of the phosphotransferase system, was inactivated, resulting in enhanced consumption of glucose. However, the surplus in substrate uptake was not converted to l-valine; instead, the formation of up to 289 μM xylulose was observed for the first time in C. glutamicum. As an alternative to the genetic engineering solution, a straightforward process engineering approach is proposed. Acetate limitation resulted in a more efficient use of acetate as cosubstrate, shown by an increased biomass yield Y _X/Ac and improved l-valine formation.     

Microfluidic study of effects of flow velocity and nutrient concentration on biofilm accumulation and adhesive strength in the flowing and no-flowing microchannels

Journal of Industrial Microbiology & Biotechnology - Biofilm accumulation in porous media can cause pore plugging and change many of the physical properties of porous media. Engineering...     

Design,synthesis, and in vitro evaluation of BP-1-102 analogs with modified hydrophobic fragments for STAT3 inhibition

Twelve novel analogs of STAT3 inhibitor BP-1-102 were designed and synthesised with the aim to modify hydrophobic fragments of the molecules that are important for interaction with the STAT3 SH2 domain. The cytotoxic activity of the reference and novel compounds was evaluated using several human and two mouse cancer cell lines. BP-1-102 and its two analogs emerged as effective cytotoxic agents and were further tested in additional six human and two murine cancer cell lines, in all of which they manifested the cytotoxic effect in a micromolar range. Reference compound S3I-201.1066 was found ineffective in all tested cell lines, in contrast to formerly published data. The ability of selected BP-1-102 analogs to induce apoptosis and inhibition of STAT3 receptor-mediated phosphorylation was confirmed. The structure–activity relationship confirmed a demand for two hydrophobic substituents, i.e. the pentafluorophenyl moiety and another spatially bulky moiety, for effective cytotoxic activity and STAT3 inhibition.     

Growth and development of the human infant breast.

Seventy-two samples of infant breasts, aged from newborn to 2 years, were collected at necropsy. Whole-mount preparations and histological sections were made. A system of classification was devised to study the extent of the structural development of the ductal system (morphological types I, II, and III) and the functional differentiation of the lining epithelium (functional stages I to V). There was no correlation between the age of the infant and the type of development of the ductal system. In contrast, the epithelial differentiation followed a chronological pattern, starting with secretory changes and apparently going through a period characterized by apocrine metaplasia before post-secretory involution. These epithelial changes were not associated with the morphological type of the ductal system. There were no distinguishing features between the breasts from the two sexes. Immunoperoxidase staining for actin and kappa-casein was carried out to study the myoepithelial cells and secretory cells, respectively. Myoepithelial cells were present at all stages and prominent staining for casein was observed up to 2 months of age. Embryonic-type adipose tissue was seen in 7 cases, in one of which it was associated closely with the developing ductal system. Extramedullary hematopoiesis was observed in the periductal connective tissue until 4 months of age. This paper describes the most extensive anatomical and histological study of the human infant breast to date and lays the foundation for a detailed study of the epithelial and stromal changes that take place during human breast development.     

Human thymic epithelial cells are frequently transformed by retroviral vectors encoding simian virus 40.

The thymic microenvironment contains a mixture of phenotypically distinct epithelial cells of varied functions, some of which are unknown. In an attempt to understand their relevance to T cell differentiation in the thymus, human thymic epithelial cell clones from both fetal (SM3-SM5) and postnatal (SM6) thymus were produced by using a defective recombinant retroviral vector encoding the simian virus 40 large T antigen and the neomycin resistance gene. The presence of keratins 8 and 18, desmosomes, and tonofilaments confirmed the epithelial origin of the cell strains. The cells expressed Thy-1 and HLA-Class I at high levels, showed weak-expression antigens defined by TE3B and A2B5, and low to negligible levels of the MR19-defined molecule. When compared with the phenotype of thymic epithelial cells in situ, the cell strains appear to be derived from neuroendocrine components in the outer cortical region of the human thymus. The use of retroviral vectors to transform human thymic epithelium was considerably more efficient than transfection with a plasmid carrying the origin of replication-defective SV40 large T gene. In the latter case, only two cell strains with subcapsular epithelial phenotypes were derived from fetal thymus. With the retroviral vectors, epithelial cell strains could, for the first time, be generated from human postnatal thymus as well as from fetal thymus.     

A chemical screen for modulators of mRNA translation identifies a distinct mechanism of toxicity for sphingosine kinase inhibitors

We here conducted an image-based chemical screen to evaluate how medically approved drugs, as well as drugs that are currently under development, influence overall translation levels. None of the compounds up-regulated translation, which could be due to the screen being performed in cancer cells grown in full media where translation is already present at very high levels. Regarding translation down-regulators, and consistent with current knowledge, inhibitors of the mechanistic target of rapamycin (mTOR) signaling pathway were the most represented class. In addition, we identified that inhibitors of sphingosine kinases (SPHKs) also reduce mRNA translation levels independently of mTOR. Mechanistically, this is explained by an effect of the compounds on the membranes of the endoplasmic reticulum (ER), which activates the integrated stress response (ISR) and contributes to the toxicity of SPHK inhibitors. Surprisingly, the toxicity and activation of the ISR triggered by 2 independent SPHK inhibitors, SKI-II and ABC294640, the latter in clinical trials, are also observed in cells lacking SPHK1 and SPHK2. In summary, our study provides a useful resource on the effects of medically used drugs on translation, identified compounds capable of reducing translation independently of mTOR and has revealed that the cytotoxic properties of SPHK inhibitors being developed as anticancer agents are independent of SPHKs.

A chemical screen to evaluate how 4100 drugs modulate translation rates confirms mTOR as the main pathway regulating translation and reveals that sphingosine kinase inhibitors downregulate translation via activation of the ER-stress response. Sphingosine kinase inhibitors, including one in clinical trials, activate stress responses and kill cells independently of the cognate target.