UPF1, a Conserved Nonsense-Mediated mRNA Decay Factor, Regulates Cyst Wall Protein Transcripts in Giardia lamblia

The Giardia lamblia cyst wall is required for survival outside the host and infection. Three cyst wall protein (cwp) genes identified to date are highly up-regulated during encystation. However, little is known of the molecular mechanisms governing their gene regulation. Messenger RNAs containing premature stop codons are rapidly degraded by a nonsense-mediated mRNA decay (NMD) system to avoid production of non-functional proteins. In addition to RNA surveillance, NMD also regulates thousands of naturally occurring transcripts through a variety of mechanisms. It is interesting to know the NMD pathway in the primitive eukaryotes. Previously, we have found that the giardial homologue of a conserved NMD factor, UPF1, may be functionally conserved and involved in NMD and in preventing nonsense suppression. In this study, we tested the hypothesis that NMD factors can regulate some naturally occurring transcripts in G. lamblia. We found that overexpression of UPF1 resulted in a significant decrease of the levels of CWP1 and cyst formation and of the endogenous cwp1-3, and myb2 mRNA levels and stability. This indicates that NMD could contribute to the regulation of the cwp1-3 and myb2 transcripts, which are key to G. lamblia differentiation into cyst. Interestingly, we also found that UPF1 may be involved in regulation of eight other endogenous genes, including up-regulation of the translation elongation factor gene, whose product increases translation which is required for NMD. Our results indicate that NMD factor could contribute to the regulation of not only nonsense containing mRNAs, but also mRNAs of the key encystation-induced genes and other endogenous genes in the early-diverging eukaryote, G. lamblia

Amyloid-Beta (Aβ) D7H Mutation Increases Oligomeric Aβ42 and Alters Properties of Aβ-Zinc/Copper Assemblies

Amyloid precursor protein (APP) mutations associated with familial Alzheimer's disease (AD) usually lead to increases in amyloid β-protein (Aβ) levels or aggregation. Here, we identified a novel APP mutation, located within the Aβ sequence (AβD7H), in a Taiwanese family with early onset AD and explored the pathogenicity of this mutation. Cellular and biochemical analysis reveal that this mutation increased Aβ production, Aβ42/40 ratio and prolonged Aβ42 oligomer state with higher neurotoxicity. Because the D7H mutant Aβ has an additional metal ion-coordinating residue, histidine, we speculate that this mutation may promote susceptibility of Aβ to ion. When co-incubated with Zn2+ or Cu2+, AβD7H aggregated into low molecular weight oligomers. Together, the D7H mutation could contribute to AD pathology through a “double punch” effect on elevating both Aβ production and oligomerization. Although the pathogenic nature of this mutation needs further confirmation, our findings suggest that the Aβ N-terminal region potentially modulates APP processing and Aβ aggregation, and further provides a genetic indication of the importance of Zn2+ and Cu2+ in the etiology of AD

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

N,N′-Bis(pyridin-3-yl)oxamide

The title molecule, C12H10N4O2, located about an inversion centre, is roughly planar, with an r.m.s. deviation from the least-squares plane of all non-H atoms of 0.019 Å. In the crystal, N—H...N hydrogen bonds between the amide N—H group and the pyridine N atom connect the molecules into a corrugated layer parallel to (10-1)

Structural directing roles of isomeric phenylenediacetate ligands in the formation of coordination networks based on flexible, N,N'-di(3-pyridyl)suberoamide

Reactions of the flexible N,N′-di(3-pyridyl)suberoamide (L) with Cu(ii) salts in the presence of the isomeric phenylenediacetic acids under hydrothermal conditions afforded three new coordination networks, {[Cu(L)(1,2-pda)]·H2O}n (1,2-H2pda = 1,2-phen

Direct and Label-Free Determination of Human Glycated Hemoglobin Levels Using Bacteriorhodopsin as the Biosensor Transducer

Glycated hemoglobin (HbA1c) levels are an important index for the diagnosis and long-term control of diabetes. This study is the first to use a direct and label-free photoelectric biosensor to determine HbA1c using bacteriorhodopsin-embedded purple membranes (PM) as a transducer. A biotinylated PM (b-PM) coated electrode that is layered with protein A-oriented antibodies against hemoglobin (Hb) readily captures non-glycated Hb (HbA0) and generates less photocurrent. The spectra of bacteriorhodopsin and Hb overlap so the photocurrent is reduced because of the partial absorption of the incident light by the captured Hb molecules. Two HbA0 and HbA1c aptasensors that are prepared by conjugating specific aptamers on b-PM coated electrodes single-step detect HbA0 and HbA1c in 15 min, without cross reactivity, with detection limits of ≤0.1 μg/mL and a dynamic range of 0.1–100 μg/mL. Both aptasensors exhibit high selectivity and long-term stability. For the clinical samples, HbA0 concentrations and HbA1c levels that are measured with aptasensors correlate well with total Hb concentrations and the HbA1c levels that are determined using standard methods (correlation gradient = 0.915 ± 0.004 and 0.981 ± 0.001, respectively). The use of these aptasensors for diabetes care is demonstrated

Self-Catenated Coordination Polymers Involving Bis-pyridyl-bis-amide

The synthesis, structures, and properties of five new coordination polymers containing bis-pyridyl-bis-amide and polycarboxylate ligands, {[Cd­(1,2-bdc)­(<b>L</b>^<b>1</b>)­(H₂O)]·H₂O}_<i>n</i> [1,2-H₂bdc = benzene-1,2-dicarboxylic acid; <b>L</b>^<b>1</b> = <i>N,N</i>′-di­(3-pyridyl)­suberoamide] (<b>1</b>), {[Cd₂(1,3-bdc)₂(<b>L</b>^<b>1</b>)₃]·8H₂O}_<i>n</i> [1,3-H₂bdc = benzene-1,3-dicarboxylic acid] (<b>2</b>), [Cd­(1,4-bdc)­(<b>L</b>^<b>1</b>)­(H₂O)₂]_<i>n</i> [1,4-H₂bdc = benzene-1,4-dicarboxylic acid] (<b>3</b>), {[Cd₂(1,2,3-Hbtc)₂(<b>L</b>^<b>1</b>)₃]·4H₂O}_<i>n</i> [1,2,3-H₃btc = 1,2,3-benzene-tricarboxylic acid] (<b>4</b>), and [Cd­(1,4-pda)­(<b>L</b>^<b>2</b>)]_<i>n</i> [<b>L</b>^<b>2</b> = <i>N,N</i>′-di­(3-pyridyl)­adipoamide; 1,4-H₂pda = 1,4-phenylenediacetic acid] (<b>5</b>), are reported, which have been characterized by single crystal X-ray diffraction. Complex <b>1</b> forms a looplike 1D chain, whereas <b>2</b> and <b>4</b> show 3D frameworks with a new self-catenated net with point symbol (6⁵·8). Complex <b>3</b> displays a 2D layer with the <b>sql</b> topology and <b>5</b> displays a rare 8T17 self-catenated net. The dimensionality and structural types of <b>1</b>–<b>3</b> can be adjusted by varying the ligand isomerism of the dicarboxylate ligands

Aβ morphology in the presence or absence of metal ions was revealed by TEM.

<p>Lyophilized Aβ was prepared in HFIP-DMSO. After 264–312 h of incubation in either the presence or absence of Zn²⁺ or Cu²⁺, the Aβ samples were stained by 2% uranyl acetate and monitored by TEM. In the presence of ions, the Aβ_D7H peptides were predominantly amorphous morphology but not protofibrils as Aβ_wt. Scale bar: 200 nm.</p