Three structural representatives of the PF06855 protein domain family from Staphyloccocus aureus and Bacillus subtilis have SAM domain-like folds and different functions

Protein domain family PF06855 (DUF1250) is a family of small domains of unknown function found only in bacteria, and mostly in the order Bacillales and Lactobacillales. Here we describe the solution NMR or X-ray crystal structures of three representatives of this domain family, MW0776 and MW1311 from Staphyloccocus aureus and yozE from Bacillus subtilis. All three proteins adopt a four-helix motif similar to sterile alpha motif (SAM) domains. Phylogenetic analysis classifies MW1311 and yozE as functionally equivalent proteins of the UPF0346 family of unknown function, but excludes MW0776, which likely has a different biological function. Our structural characterization of the three domains supports this separation of function. The structures of MW0776, MW1311, and yozE constitute the first structural representatives from this protein domain family.     

Novel sesquiterpene and copyrine alkaloids from Anaxagorea javanica Blume

A novel sesquiterpene, nordine (or 2,2,9-trimethyl-5-methylene-12-oxa-bicyclo[6.3.1]dodecane-4,9-diol) (1) and two new copyrine alkaloids of eupolauridine skeleton, viz., 4,11-dimethoxyeupolauridine (2) and 2-methoxy-3-hydroxyeupolauridine (3), along with five alkaloidal (4–8) and nine non-alkaloidal compounds (9–17), have been isolated from the bark and roots of Anaxagorea javanica Blume. The structures of the new constituents were deduced by NMR spectral data. Compounds 6 and 7 displayed moderate inhibition of NO production with IC₅₀ values of 18.2 and 32.3 μg/ml respectively.     

Identification of Stk25 as a genetic modifier of Tau phosphorylation in Dab1-mutant mice

Hyperphosphorylation of the microtubule binding protein Tau is a feature of a number of neurodegenerative diseases, including Alzheimer's disease. Tau is hyperphosphorylated in the hippocampus of dab1-null mice in a strain-dependent manner; however, it has not been clear if the Tau phosphorylation phenotype is a secondary effect of the morbidity of these mutants. The dab1 gene encodes a docking protein that is required for normal brain lamination and dendritogenesis as part of the Reelin signaling pathway. We show that dab1 gene inactivation after brain development leads to Tau hyperphosphorylation in anatomically normal mice. Genomic regions that regulate the phospho Tau phenotype in dab1 mutants have previously been identified. Using a microarray gene expression comparison between dab1-mutants from the high-phospho Tau expressing and low-phospho Tau expressing strains, we identified Stk25 as a differentially expressed modifier of dab1-mutant phenotypes. Stk25 knockdown reduces Tau phosphorylation in embryonic neurons. Furthermore, Stk25 regulates neuronal polarization and Golgi morphology in an antagonistic manner to Dab1. This work provides insights into the complex regulation of neuronal behavior during brain development and provides insights into the molecular cascades that regulate Tau phosphorylation.     

Comparison of identified mitral and tufted cells in freely breathing rats: II. Odor-evoked responses

Mitral and tufted cells are the 2 types of output neurons of the main olfactory bulb. They are located in distinct layers, have distinct projection patterns of their dendrites and axons, and likely have distinct relationships with the intrabulbar inhibitory circuits. They could thus be functionally distinct and process different aspects of olfactory information. To examine this possibility, we compared the odor-evoked responses of identified single units recorded in the mitral cell layer (MCL units), in the core of the external plexiform layer (not at the glomerular border tufted cells), or at the glomerular border of this layer (GB tufted cells) of the entire olfactory bulb. Differences between mitral and tufted cells were observed only when subtle aspects of the responses were explored, such as the firing rate per respiratory cycle or the distribution of firing activity along the respiratory cycle. By contrast, more clear differences were found when the 2 subtypes of tufted cells were examined separately. GB units were significantly more responsive, had significantly higher firing activity, and showed greater activity at the transition between inspiration and expiration. The projection-type tufted cells situated closer to the entrance of the olfactory bulb may thus form a distinct physiological class of output neurons and differ from mitral cells and other tufted cells in the manner of processing olfactory information.     

Erythrocyte Indices, Microminerals and Ratios, Antioxidants and Lipids in Centrum Materna Diet-Supplemented Omani Mothers

Venous (maternal) and cord blood (neonatal) samples of Omani women who had a daily supplement of Centrum Materna multivitamin and multimineral tablet throughout pregnancy were investigated at late preterm (n=37) and at term (n=37) delivery for erythrocyte indices, micromineral, antioxidant, and lipid values. Hemoglobin (Hb), hematocrit (HCT), mean cell volume (MCV), red cell distribution width (RDW), copper (Cu), zinc (Zn), ceruloplasmin, erythrocyte Cu-Zn superoxide dismutase (Cu-Zn SOD), cholesterol, apolipoprotein (apo) A-I and apo B were measured by appropriate analytical systems. Cu/zinc and Cu/ceruloplasmin ratios were calculated. The erythrocyte indices were normal in neonatal blood but showed borderline anemia in maternal blood of both groups. There were significantly decreased values of Cu (P=0.012), Zn (P=0.001), apo A-I (P=0.029), and Cu/ceruloplasmin ratio (P=0.032) in late preterm compared to term mothers. Significantly decreased values of Cu (P=0.003), ceruloplasmin (P<0.0001), apo A-I (P=0.024), and Cu/Zn ratio (P=007) were observed in late preterm relative to term neonates. Late preterm mothers were significantly younger (P=0.027) than term mothers. Maternal age correlated positively with apo A-I (r=0.424, P=0.012) and negatively with Cu/Zn ratio (r=-0.353, P=0.040). The findings suggest that with daily dietary Centrum Materna supplementation throughout pregnancy, hematological indices were maintained within normal in mothers and neonates, but the levels of microminerals and micromineral ratios were subnormal in late preterm mothers and their neonates.     

Impact of dust exposure on mixed bacterial cultures and during eukaryotic cell co-culture infections

On a daily basis, humans, and their colonizing microbiome, are exposed to both indoor and outdoor dust, containing both deleterious organic and inorganic contaminants, through dermal contact, inhalation, and ingestion. Recent studies evaluating the dust exposure responses of opportunistic pathogens, such as Escherichia coli and Pseudomonas aeruginosa, revealed significant increases in biofilm formation following dust exposure. In this study, the effects of dust exposure on mixed bacterial cultures as well as HT-29 co-cultures were evaluated. As it was observed in pure, single bacterial cultures earlier, neither indoor nor outdoor dust exposure (at concentrations of 100 μg/mL) influenced the growth of mixed bacterial liquid cultures. However, when in paired mixed cultures, dust exposure increased sensitivity to oxidative stress and significantly enhanced biofilm formation (outdoor dust). More specifically, mixed cultures (E. coli-Klebsiella pneumoniae, K. pneumoniae-P. aeruginosa, and E. coli-P. aeruginosa) exhibited increased sensitivity to 20 and 50 mM of H2O2 in comparison to their pure, single bacterial culture counterparts and significantly enhanced biofilm production for each mixed culture. Finally, bacterial proliferation during a eukaryotic gut cell (HT29) co-culture was significantly more robust for both K. pneumoniae and P. aeruginosa when exposed to both house and road dust; however, E. coli only experienced significantly enhanced proliferation, in HT29 co-culture, when exposed to road dust. Taken together, our findings demonstrate that bacteria respond to dust exposure differently when in the presence of multiple bacterial species or when in the presence of human gut epithelial cells, than when grown in isolation.     

A genomic sequence analysis of the mouse and human microtubule-associated protein tau

Microtubule associated protein tau (MAPT) encodes the microtubule associated protein tau, the primary component of neurofibrillary tangles found in Alzheimer's disease and other neurodegenerative disorders. Mutations in the coding and intronic sequences of MAPT cause autosomal dominant frontotemporal dementia (FTDP-17). MAPT is also a candidate gene for progressive supranuclear palsy and hereditary dysphagic dementia. A human PAC (201 kb) and a mouse BAC (161 kb) containing the entire MAPT and Mtapt genes, respectively, were identified and sequenced. Comparative DNA sequence analysis revealed over 100 conserved non-repeat potential cis-acting regulatory sequences in or close to MAPT. Those islands with greater than 67% nucleotide identity range in size from 20 to greater than 1700 nucleotides. Over 90 single nucleotide polymorphisms were identified in MAPT that are candidate susceptibility alleles for neurodegenerative disease. The 5′ and 3′ flanking genes for MAPT are the corticotrophin-releasing factor receptor (CRFR) gene and KIAA1267, a gene of unknown function expressed in brain. Received: 1 April 2001 / Accepted: 20 April 2001     

Biotechnology: Genetic improvement of sorghum (Sorghum bicolor (L.) Moench)

Summary This report reviews the contributions to the improvement of sorghum (Sorghum bicolor (L.) Moench) through traditional approaches with emphasis on the application of biotechnological methods. Strategies include breeding for higher yield, improved grain quality, and biotic and abiotic stress tolerance. Hybrid development and polyploidy breeding are also discussed. Plant breeders, working in concert with biotechnologists, have developed new powerful tools for plant genetic manipulation and genotype evaluation that will significantly improve the efficiency of plant breeding. Improving sorghum through biotechnology is the latest in a long series of technologies that have been applied to this crop. Five basic tools of technology have been developed for sorghum improvement: (1) in vitro protocols for efficient plant regeneration; (2) molecular markers; (3) gene identification and cloning; (4) genetic engineering and gene transfer technology to integrate desirable traits into the sorghum genome; and (5) genomics and germplasm databases. Reports on studies involving the problems, progress, and prospects for utilizing the biotechnological methods for sorghum improvement are discussed.     

Plant genetic engineering to improve biomass characteristics for biofuels

Currently, most ethanol produced in the United States is derived from maize kernel, at levels in excess of four billion gallons per year. Plant lignocellulosic biomass is renewable, cheap and globally available at 10-50 billion tons per year. At present, plant biomass is converted to fermentable sugars for the production of biofuels using pretreatment processes that disrupt the lignocellulose and remove the lignin, thus allowing the access of microbial enzymes for cellulose deconstruction. Both the pretreatments and the production of enzymes in microbial tanks are expensive. Recent advances in plant genetic engineering could reduce biomass conversion costs by developing crop varieties with less lignin, crops that self-produce cellulase enzymes for cellulose degradation and ligninase enzymes for lignin degradation, or plants that have increased cellulose or an overall biomass yield.     

In vivo cervical cancer growth inhibition by genetically engineered cytotoxic T cells

Purpose: The CD44 v7/8 splice variant that is frequently expressed in cervical carcinoma and rarely expressed in normal tissues displays promising properties as a target antigen for cancer immune therapy. In this study, cytotoxic T lymphocytes (CTLs) were genetically engineered to gain CD44v7/8 target specificity. Methods: Clone 96 (CI96), an established murine cytotoxic T-cell line, and naïve murine T cells were retrovirally transduced with a fusion gene construct encoding for the single chain fragment scFv of the monoclonal antibody VFF17 and for the chain of the T-cell receptor (TCR). The therapeutic potential of genetically engineered T cells was tested in vitro and in vivo. Results: Surface expression of the chimeric TCR on infected Cl96 and naïve T cells was shown by FACS analysis. CD44v7/8-positive target cells were efficiently lysed by transduced Cl96 and naïve T cells, demonstrating the functionality and specificity of the chimeric TCR. In a xenograft BALB/c mouse model, efficient growth retardation of CD44v7/8-positive tumours was mediated by genetically engineered Cl96(VFF17)cyYZ cells. Conclusions: We were able to reprogramme the target specificity of recombinant Cl96 and naïve CTLs resulting in efficient cytolysis of CD44v7/8-positive cervical cancer cells. High transduction rates and the specific cytolysis of CD44v7/8-redirected CTLs are promising tools for an immune gene therapy approach for advanced cervical cancer.Abbreviations Ab Antibody - CTL Cytolytic T lymphocyte - mAb Monoclonal antibody - TCR T-cell receptor