DNA hybridization detection with organic thin film transistors: toward fast and disposable DNA microarray chips

We demonstrate a novel DNA hybridization detection method with organic thin film transistors. DNA molecules are immobilized directly on the surface of organic semiconductors, producing an unambiguous doping-induced threshold voltage shift upon hybridization. With these shifts, single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA) are differentiated successfully. This method is expected to result in higher sensitivity than the main competitive technology, ISFET-based sensors because of the direct exposure of DNA molecules to sensitive layers. Factors that influence sensor sensitivity have been analyzed and optimum conditions have been determined using statistically designed experiments. Under the optimum conditions, the maximum difference between saturation current ratios caused by ssDNA and dsDNA reaches as high as 70%. In order to make DNA detection fast, we also demonstrate rapid on-chip electrically enhanced hybridization using the TFTs. These technologies together will enable the realization of disposable, rapid-turnaround tools for field-deployable genomic diagnosis.     

Immunocontraceptive potential of recombinantly expressed minimized chicken riboflavin carrier protein (mini-RCP) in rodents

Chicken riboflavin carrier protein (RCP; 219 AA) harbours four linear epitopes, constituted by the peptide residues 3-23, 64-83, 130-147 and 200-219. Antibodies to these sequences bioneutralize maternal RCP and provide protection from pregnancy in rodents. In order to overcome the major histocompatibility complex-dependent variability in immune response often encountered with use of single peptides for vaccination in genetically outbred populations, we have assembled a novel synthetic gene, incorporating in tandem the nucleotide sequences coding for all the four neutralizing epitopes of chicken RCP and expressed in Escherichia coli. The gene product, mini-RCP has been characterized for its immunogenic properties and contraceptive potential in rodents. Immunization of rabbits and rats led to generation of antibodies against individual peptide components, as determined by enzyme-linked-immunosorbent assay (ELISA). However, immunized rats carried pregnancy to term and delivered healthy offsprings. Antisera from these rats exhibited decreased affinity of binding to the native protein. These findings suggest that the prospects of covalently-linked epitope peptides need to be cautiously evaluated during the design and development of peptide-based vaccines.     

Ribosomal protein L35: identification in spinach chloroplasts and isolation of a cDNA clone encoding its cytoplasmic precursor

We describe the isolation of spinach chloroplast ribosomal protein L35 and characterization of a cDNA clone encoding its cytoplasmic precursor. This protein was only recently identified in ribosomes, but the sequences of four L35 genes have now been reported and confirm its presence in eubacteria, chloroplasts, and cyanelles. Using N-terminal sequence data, oligonucleotides were designed and a cDNA library was screened. The nucleotide sequence of the cDNA clones shows that the spinach L35 protein is encoded as a precursor of 159 residues, comprising a mature protein of 73 residues and a transit peptide of 86 residues. The cleavage site for forming the mature protein is deduced to be Thr-Val-Phe-Ala decreases Ala-Lys-Gly-Tyr. The L35 protein in the photosynthetic organelle of the protozoan Cyanophora paradoxa is encoded in the organelle DNA [Bryant & Stirewalt (1990) FEBS Lett. 259, 273-280]. The corresponding gene has not been found in the chloroplast DNA of a lower plant (liverwort) and two higher plants. Our results demonstrate that the L35 protein in a higher plant (spinach) is encoded in the nucleus. This finding, in light of the endosymbiont hypothesis, suggests an organelle to nucleus transfer of the L35 gene at the evolutionary beginnings of land plants.     

Design and Synthesis of 1-Substituted-4-(4-Nitrophenyl)-[1,2,4]triazolo[4,3-a]quinazolin-5(4H)-ones as a New Class of Antihistaminic Agents

Russian Journal of Bioorganic Chemistry - Some new 1-substituted-4-(4-nitrophenyl)-[1,2,4]triazolo[4,3-a]quinazolin-5(4H)-ones were synthesized and screened for their H1-antihistaminic activity....     

The amino and carboxyl termini of perilipin a facilitate the storage of triacylglycerols

Perilipin A is the most abundant lipid droplet-associated protein in adipocytes and serves important functions in regulating triacylglycerol levels by reducing rates of basal lipolysis and facilitating hormonally stimulated lipolysis. We have previously shown that the central region of perilipin A targets and anchors it to lipid droplets, at least in part via three moderately hydrophobic sequences that embed the protein into the hydrophobic core of the droplet. The current study examines the roles of the amino and carboxyl termini of perilipin A in facilitating triacylglycerol storage. Amino- and carboxyl-terminal truncation mutations of mouse perilipin A were stably expressed in 3T3-L1 preadipocytes, which lack perilipins. Triacylglycerol content of the cells was quantified as a measure of perilipin function and was compared with that of cells expressing full-length perilipin A or control cells lacking perilipins. The amino-terminal sequence between amino acids 122 and 222, including four 10-11-amino acid sequences predicted to form amphipathic beta-strands and a consensus site for cAMP-dependent protein kinase, and the carboxyl terminus of 112 amino acids that is unique to perilipin A were critical to facilitate triacylglycerol storage. The precocious expression of full-length perilipin A in 3T3-L1 preadipocytes aided more rapid storage of triacylglycerol during adipose differentiation. By contrast, the expression of highly truncated amino- or carboxyl-terminal mutations of perilipin failed to serve a dominant negative function in lowering triacylglycerol storage during adipose differentiation. We conclude that the amino and carboxyl termini are critical to the function of perilipin A in facilitating triacylglycerol storage.     

A new record of monogeneric family Vietnamellidae (Insecta: Ephemeroptera) from India

A new record of monogeneric family Vietnamellidae (Insecta: Ephemeroptera) is established for India with Vietnamella sp. A described based on the larvae from Arunachal Pradesh, India. This species can be distinguished from other known species of this genus in the larval stage by the following combination of characters: (i) outer pair of projections in head large and stout, triangular, cone-shaped with serrated spines; (ii) posterolateral angles of abdominal terga 2–9 extended into sharp projections; (iii) caudal filaments pale yellowish brown with dense lateral setae on inner and outer margins of middle part; (iv) femora of mid- and hind-legs broader; and (v) second segment of the maxillary palpi shorter than first segment.     

Stress-dependent Proteolytic Processing of the Actin Assembly Protein Lsb1 Modulates a Yeast Prion

Yeast prions are self-propagating amyloid-like aggregates of Q/N-rich protein that confer heritable traits and provide a model of mammalian amyloidoses. [PSI⁺] is a prion isoform of the translation termination factor Sup35. Propagation of [PSI⁺] during cell division under normal conditions and during the recovery from damaging environmental stress depends on cellular chaperones and is influenced by ubiquitin proteolysis and the actin cytoskeleton. The paralogous yeast proteins Lsb1 and Lsb2 bind the actin assembly protein Las17 (a yeast homolog of human Wiskott-Aldrich syndrome protein) and participate in the endocytic pathway. Lsb2 was shown to modulate maintenance of [PSI⁺] during and after heat shock. Here, we demonstrate that Lsb1 also regulates maintenance of the Sup35 prion during and after heat shock. These data point to the involvement of Lsb proteins in the partitioning of protein aggregates in stressed cells. Lsb1 abundance and cycling between actin patches, endoplasmic reticulum, and cytosol is regulated by the Guided Entry of Tail-anchored proteins pathway and Rsp5-dependent ubiquitination. Heat shock-induced proteolytic processing of Lsb1 is crucial for prion maintenance during stress. Our findings identify Lsb1 as another component of a tightly regulated pathway controlling protein aggregation in changing environments.     

Deletion of Serum Amyloid A3 Improves High Fat High Sucrose Diet-Induced Adipose Tissue Inflammation and Hyperlipidemia in Female Mice

Serum amyloid A (SAA) increases in response to acute inflammatory stimuli and is modestly and chronically elevated in obesity. SAA3, an inducible form of SAA, is highly expressed in adipose tissue in obese mice where it promotes monocyte chemotaxis, providing a mechanism for the macrophage accumulation that occurs with adipose tissue expansion in obesity. Humans do not express functional SAA3 protein, but instead express SAA1 and SAA2 in hepatic as well as extrahepatic tissues, making it difficult to distinguish between liver and adipose tissue-specific SAA effects. SAA3 does not circulate in plasma, but may exert local effects that impact systemic inflammation. We tested the hypothesis that SAA3 contributes to chronic systemic inflammation and adipose tissue macrophage accumulation in obesity using mice deficient for Saa3 (Saa3 ^−/−). Mice were rendered obese by feeding a pro-inflammatory high fat, high sucrose diet with added cholesterol (HFHSC). Both male and female Saa3 ^−/− mice gained less weight on the HFHSC diet compared to Saa3^+/+ littermate controls, with no differences in body composition or resting metabolism. Female Saa3 ^−/− mice, but not males, had reduced HFHSC diet-induced adipose tissue inflammation and macrophage content. Both male and female Saa3 ^−/− mice had reduced liver Saa1 and Saa2 expression in association with reduced plasma SAA. Additionally, female Saa3 ^−/− mice, but not males, showed improved plasma cholesterol, triglycerides, and lipoprotein profiles, with no changes in glucose metabolism. Taken together, these results suggest that the absence of Saa3 attenuates liver-specific SAA (i.e., SAA1/2) secretion into plasma and blunts weight gain induced by an obesogenic diet. Furthermore, adipose tissue-specific inflammation and macrophage accumulation are attenuated in female Saa3 ^−/− mice, suggesting a novel sexually dimorphic role for this protein. These results also suggest that Saa3 influences liver-specific SAA1/2 expression, and that SAA3 could play a larger role in the acute phase response than previously thought.     

Microbial Communities in Chesapeake & Ohio Canal National Historical Park and Their Function as Indicators of Water Quality

Abstract

The objective of this study was to establish meaningful relationships between prokaryotic community profiles and water quality parameters in different water bodies (spring, stream, cave, and mine) in the middle reach of the Chesapeake & Ohio Canal National Historic Park (C&O), Maryland. The microbial profiles in the water samples were determined using metagenomic analysis. The relationships between microbial phylogenetic profiles and water quality parameters were investigated using principal component analysis (PCA) and redundancy analysis (RDA). The most abundant phyla identified in most samples were Proteobacteria (55.4%), Bacteroidetes (12.3%), Actinobacteria (10.6%), Firmicutes (2.4%), Planktomycetes (1.8%), Verrucomicrobia (1.5%), Chloroflexi (1.5%), and Acidobacteria (1.3%), which are major bacterial and archaeal groups typically observed in natural freshwater environments. PCA showed that water chemistry was determined primarily by the geology of the site and the type of water source (i.e., spring, stream, cave, or mine). Most samples located in carbonate formations correlated with high alkalinity, inorganic carbon, and calcium, representing the typical karstic geochemistry. RDA shows that pH, electrical conductivity, temperature and nutrients including nitrate, phosphate, and sulfate, were significant determinants of the microbial ecology.