Monoclonal antibodies specific for the core protein of the beta-subunit of the gastric proton pump (H+/K+ ATPase). An autoantigen targetted in pernicious anaemia.

The gastric H+/K(+)-transporting adenosine triphosphatase (H+/K+ ATPase) (proton pump) consists of a catalytic alpha-subunit and a recently proposed 60-90-kDa glycoprotein beta-subunit. Using dog gastric membranes as the antigen, we have produced two murine monoclonal antibodies, 4F11 (IgG1) and 3A6 (IgA), which are specific for the 60-90-kDa glycoprotein. The monoclonal antibodies (1) specifically stained the cytoplasm of unfixed and formalin-fixed dog gastric parietal cells; (2) specifically reacted by ELISA with gastric tubulovesicular membranes; (3) recognised epitopes located on the luminal face of parietal cell tubulovesicular membranes, the site of the proton pump, by immunogold electron microscopy; (4) immunoblotted a 60-90-kDa molecule from tubulovesicular membranes and a 35-kDa component from peptide N-glycosidase-F-treated membrane extracts; (5) immunoblotted the 60-90-kDa parietal cell autoantigen associated with autoimmune gastritis and pernicious anemia, purified by chromatography on parietal cell autoantibody- or tomato-lectin-Sepharose 4B affinity columns, and the 35-kDa protein core of this autoantigen; this autoantigen has amino acid sequence similarity to the beta-subunit of the related Na+/K(+)-transporting adenosine triphosphatase (Na+/K+ ATPase) [Toh et al. (1990) Proc. Natl Acad. Sci. 87, 6418-6422]; (6) co-precipitated a molecule of 95 kDa with the 60-90-kDa molecule from 125I-labelled detergent extracts of dog tubulovesicular membranes; and (7) co-purified the catalytic alpha-subunit of the H+/K+ ATPase with the 60-90-kDa molecule by immunoaffinity chromatography of tubulovesicular membrane extracts on a monoclonal antibody 3A6-Sepharose 4B column, indicating a physical association between the two molecules. These results provide further evidence that the 60-90-kDa glycoprotein is the beta-subunit of the gastric H+/K+ ATPase. We conclude that the monoclonal antibodies specifically recognise luminal epitopes on the 35-kDa core protein of the 60-90-kDa beta-subunit of the gastric proton pump, a major target molecule in autoimmune gastritis and pernicious anaemia. These monoclonal antibodies will be valuable probes to study the structure and function of this associated beta-subunit, as well as the ontogeny of the gastric proton pump.     

Human long-chain acyl-CoA synthetase: structure and chromosomal location.

A complementary DNA clone encoding the entire human long-chain acyl-CoA synthetase was isolated and the total 698-amino acid sequence was deduced. The amino acid sequence of human long-chain acyl-CoA synthetase shows 84.9% identity to that of rat long-chain acyl-CoA synthetase. The nucleotide sequences of the protein coding regions between human and rat long-chain acyl-CoA synthetase mRNAs are highly conserved (85.6%), whereas those of the 3' untranslated regions are less conserved (72%). The location of the human long-chain acyl-CoA synthetase gene was identified on chromosome 4 by spot hybridization of flow-sorted chromosomes. Computer-assisted homology search revealed a significant similarity of the enzyme with the enzymes of the luciferase family. Based on this similarity, the structure of human long-chain acyl-CoA synthetase can be divided into five domains: the N-terminus, two domains similar to those in enzymes of the luciferase family, a long gap region between the similar domains and the C-terminus.     

Impact of Vitamin D Replacement on Markers of Glucose Metabolism and Cardio-Metabolic Risk in Women with Former Gestational Diabetes—A Double-Blind,Randomized Controlled Trial

Gestational Diabetes Mellitus (GDM) and vitamin D deficiency are related to insulin resistance and impaired beta cell function, with heightened risk for future development of diabetes. We evaluated the impact of vitamin D supplementation on markers of glucose metabolism and cardio metabolic risk in Asian women with former GDM and hypovitaminosis D. In this double blind, randomized controlled trial, 26 participants were randomized to receive either daily 4000 IU vitamin D3 or placebo capsules. 75g Oral Glucose Tolerance Test (OGTT) and biochemistry profiles were performed at baseline and 6 month visits. Mathematical models, using serial glucose, insulin and C peptide measurements from OGTT, were employed to calculate insulin sensitivity and beta cell function. Thirty three (76%) women with former GDM screened had vitamin D level of <50 nmol/L at baseline. Supplementation, when compared with placebo, resulted in increased vitamin D level (+51.1 nmol/L vs 0.2 nmol/L, p<0.001) and increased fasting insulin (+20% vs 18%, p = 0.034). The vitamin D group also demonstrated a 30% improvement in disposition index and an absolute 0.2% (2 mmol/mol) reduction in HbA_1c. There was no clear change in insulin sensitivity or markers of cardio metabolic risk. This study highlighted high prevalence of vitamin D deficiency among Asian women with former GDM. Six months supplementation with 4000 IU of vitamin D3 safely restored the vitamin D level, improved basal pancreatic beta-cell function and ameliorated the metabolic state. There was no effect on markers of cardio metabolic risk. Further mechanistic studies exploring the role of vitamin D supplementation on glucose homeostasis among different ethnicities may be needed to better inform future recommendations for these women with former GDM at high risk of both hypovitaminosis D and future diabetes.     

Similarities and seasonal variations in bacterial communities from the blood of rodents and from their flea vectors

Vector-borne microbes are subject to the ecological constraints of two distinct microenvironments: that in the arthropod vector and that in the blood of its vertebrate host. Because the structure of bacterial communities in these two microenvironments may substantially affect the abundance of vector-borne microbes, it is important to understand the relationship between bacterial communities in both microenvironments and the determinants that shape them. We used pyrosequencing analyses to compare the structure of bacterial communities in Synosternus cleopatrae fleas and in the blood of their Gerbillus andersoni hosts. We also monitored the interindividual and seasonal variability in these bacterial communities by sampling the same individual wild rodents during the spring and again during the summer. We show that the bacterial communities in each sample type (blood, female flea or male flea) had a similar phylotype composition among host individuals, but exhibited seasonal variability that was not directly associated with host characteristics. The structure of bacterial communities in male fleas and in the blood of their rodent hosts was remarkably similar and was dominated by flea-borne Bartonella and Mycoplasma phylotypes. A lower abundance of flea-borne bacteria and the presence of Wolbachia phylotypes distinguished bacterial communities in female fleas from those in male fleas and in rodent blood. These results suggest that the overall abundance of a certain vector-borne microbe is more likely to be determined by the abundance of endosymbiotic bacteria in the vector, abundance of other vector-borne microbes co-occurring in the vector and in the host blood and by seasonal changes, than by host characteristics.     

Forest resistance to drought in a humid tropical dipterocarp forest in the Western Ghats of India

Droughts are linked to tree die-offs in the biodiverse humid tropics. We assessed drought response of a Dipterocarp Forest and found a marginal decrease in tree survival, indicating drought resistance. Understory and emergent species were sensitive to drought. Urgent focus is needed to understand drought impacts and plant physiological responses in South Asian forests.     

Soy‐Derived Phytochemical Genistein Modifies Chromatome Topology to Restrict Cancer Cell Proliferation

Epidemiological data indicate that human cancer risk is significantly reduced by the consumption of soy‐based foods containing the “phytoestrogen” genistein, which can signal via host cell estrogen receptors. While additional chemoprotective effects of genistein induced by epigenetic factors have also been reported, the key molecules and mechanisms involved are poorly defined. We therefore investigated genistein effects on chromatin‐bound proteins in the estrogen receptor‐deficient cell line MDA‐MB‐231 which is insensitive to phytoestrogen signaling. After exposure to low‐dose genistein for >1 month, MDA‐MB‐231 cells exhibited stable epigenetic alterations that are analyzed via partial MNase digestion and TMT‐based quantitative proteomics. 3177 chromatin‐bound proteins are identified with high confidence, including 882 molecules that displayed altered binding topology after cell conditioning with genistein. Prolonged phytochemical exposure conferred heritable changes in the binding topology of key epigenetic regulators including ATRX, SUV39H1/H2, and HP1BP3 that are preserved in untreated progeny, resulting in sustained downregulation of proliferation genes and reduced cell growth. These data indicate that soy derivative genistein exerts complex estrogen receptor‐independent effects on the epigenome likely to influence tumorigenesis by restricting cell growth.     

Complete Genome Sequence and Comparative Analysis of the Wild-type Commensal Escherichia coli Strain SE11 Isolated from a Healthy Adult

We sequenced and analyzed the genome of a commensal Escherichia coli (E. coli) strain SE11 (O152:H28) recently isolated from feces of a healthy adult and classified into E. coli phylogenetic group B1. SE11 harbored a 4.8 Mb chromosome encoding 4679 protein-coding genes and six plasmids encoding 323 protein-coding genes. None of the SE11 genes had sequence similarity to known genes encoding phage- and plasmid-borne virulence factors found in pathogenic E. coli strains. The comparative genome analysis with the laboratory strain K-12 MG1655 identified 62 poorly conserved genes between these two non-pathogenic strains and 1186 genes absent in MG1655. These genes in SE11 were mostly encoded in large insertion regions on the chromosome or in the plasmids, and were notably abundant in genes of fimbriae and autotransporters, which are cell surface appendages that largely contribute to the adherence ability of bacteria to host cells and bacterial conjugation. These data suggest that SE11 may have evolved to acquire and accumulate the functions advantageous for stable colonization of intestinal cells, and that the adhesion-associated functions are important for the commensality of E. coli in human gut habitat.Key words: Escherichia coli, commensal, human gut, genome sequencing     

Response properties of visual interneurons to motion stimuli in the praying mantis,Tenodera aridifolia

Intracellular responses of motion-sensitive visual interneurons were recorded from the lobula complex of the mantis, Tenodera aridifolia. The interneurons were divided into four classes according to the response polarity, spatial tuning, and directional selectivity. Neurons of the first class had small, medium, or large receptive fields and showed a strong excitation in response to a small-field motion such as a small square moving in any direction (SF neurons). The second class neurons showed non-directionally selective responses: an excitation to a large-field motion of gratings in any direction (ND neurons). Most ND neurons had small or medium-size receptive fields. Neurons of the third class had large receptive fields and exhibited directionally selective responses: an excitation to a large-field motion of gratings in preferred direction and an inhibition to a motion in opposite, null direction (DS neurons). The last class neurons had small receptive fields and showed inhibitory responses to a moving square and gratings (I neurons). The functional roles of these neurons in prey recognition and optomotor response were discussed.     

The contrasting roles of Th17 immunity in human health and disease

The human immune system is a tightly regulated network that protects the host from disease. An important aspect of this is the balance between pro‐inflammatory Th17 cells and anti‐inflammatory T regulatory (Treg) cells in maintaining immune homeostasis. Foxp3+ Treg are critical for sustaining immune tolerance through IL‐10 and transforming growth factor‐β while related orphan receptor‐γt+ Th17 cells promote immunopathology and auto‐inflammatory diseases through the actions of IL‐17A, IL‐21 and IL‐22. Therefore, imbalance between Treg and Th17 cells can result in serious pathology in many organs and tissues. Recently, certain IL‐17‐producing cells have been found to be protective against infectious disease, particularly in relation to extracellular bacteria such Streptococcus pneumoniae; a number of other novel IL‐17‐secreting cell populations have also been reported to protect against a variety of other pathogens. In this mini‐review, the dual roles of Treg and Th17 cells are discussed in the context of autoimmunity and infections, highlighting recent advances in the field. Development of novel strategies specifically designed to target these critical immune response pathways will become increasingly important in maintenance of human health.