Hsc66 substrate specificity is directed toward a discrete region of the iron-sulfur cluster template protein IscU

Hsc66 and Hsc20 comprise a specialized chaperone system important for the assembly of iron-sulfur clusters in Escherchia coli. Only a single substrate, the Fe/S template protein IscU, has been identified for the Hsc66/Hsc20 system, but the mechanism by which Hsc66 selectively binds IscU is unknown. We have investigated Hsc66 substrate specificity using phage display and a peptide array of IscU. Screening of a heptameric peptide phage display library revealed that Hsc66 prefers peptides with a centrally located Pro-Pro motif. Using a cellulose-bound peptide array of IscU we determined that Hsc66 interacts specifically with a region (residues 99-103, LPPVK) that is invariant among all IscU family members. A synthetic peptide (ELPPVKIHC) corresponding to IscU residues 98-106 behaves in a similar manner to native IscU, stimulating the ATPase activity of Hsc66 with similar affinity as IscU, preventing Hsc66 suppression of bovine rhodanese aggregation, and interacting with the peptide-binding domain of Hsc66. Unlike native IscU, however, the synthetic peptide is not bound by Hsc20 and does not synergistically stimulate Hsc66 ATPase activity with Hsc20. Our results indicate that Hsc66 and Hsc20 recognize distinct regions of IscU and further suggest that Hsc66 will not bind LPPVK motifs with high affinity in vivo unless they are in the context of native IscU and can be directed to Hsc66 by Hsc20

Three hydrophobic amino acids in Escherichia coli HscB make the greatest contribution to the stability of the HscB-IscU complex

<p>Abstract</p> <p>Background</p> <p>General iron-sulfur cluster biosynthesis proceeds through assembly of a transient cluster on IscU followed by its transfer to a recipient apo-protein. The efficiency of the second step is increased by the presence of HscA and HscB, but the reason behind this is poorly understood. To shed light on the function of HscB, we began a study on the nature of its interaction with IscU. Our work suggested that the binding site of IscU is in the C-terminal domain of HscB, and two different triple alanine substitutions ([L92A, M93A, F153A] and [E97A, E100A, E104A]) involving predicted binding site residues had detrimental effects on this interaction. However, the individual contribution of each substitution to the observed effect remains to be determined as well as the possible involvement of other residues in the proposed binding site.</p> <p>Results</p> <p>In the work reported here, we used isothermal titration calorimetry to characterize the affinity of single alanine HscB mutants for IscU, and subsequently confirmed our results with nuclear magnetic resonance spectroscopy. Alanine substitutions of L92, L96, and F153 severely impaired the ability of HscB to form a complex with IscU; substitutions of R87, R99, and E100 had more modest effects; and substitutions of T89, M93, E97, D103, E104, R152, K156, and S160 had only minor or no detectable effects.</p> <p>Conclusions</p> <p>Our results show that the residues of HscB most important for strong interaction with IscU include three hydrophobic residues (L92, L96, and F153); in addition, we identified a number of other residues whose side chains contribute to a lesser extent to the interaction. Our results suggest that the triple alanine substitution at HscB positions 92, 96, and 153 will destabilize the HscB-IscU complex by ΔΔ<it>G</it>_b≅ 5.7 kcal/mol, equivalent to a ≅ 15000-fold reduction in the affinity of HscB for IscU. We propose that this triple mutant could provide a more definitive test of the functional importance of the HscB-IscU interaction in vivo than those used previously that yielded inconclusive results.</p

Linear confinement without dilaton in bottom-up holography for walking technicolour

In PRD78(2008)055005 [arXiv:0805.1503 [hep-ph]] and PRD79(2009)075004 [arXiv:0809.1324 [hep-ph]], we constructed a holographic description of walking technicolour theories using both a hard- and a soft-wall model. Here, we show that the dilaton field becomes phenomenologically irrelevant for the spectrum of spin-one resonances once a term is included in the Lagrangian that mixes the Goldstone bosons and the longitudinal components of the axial vector mesons. We show how this mixing affects our previous results and we make predictions about how this description of technicolour can be tested.Comment: 7 pages, no figure

Serum microRNA array analysis identifies miR-140-3p, miR-33b-3p and miR-671-3p as potential osteoarthritis biomarkers involved in metabolic processes.

Background: MicroRNAs (miRNAs) in circulation have emerged as promising biomarkers. In this study, we aimed to identify a circulating miRNA signature for osteoarthritis (OA) patients and in combination with bioinformatics analysis to evaluate the utility of selected differentially expressed miRNAs in the serum as potential OA biomarkers. Methods: Serum samples were collected from 12 primary OA patients, and 12 healthy individuals were screened using the Agilent Human miRNA Microarray platform interrogating 2549 miRNAs. Receiver Operating Characteristic (ROC) curves were constructed to evaluate the diagnostic performance of the deregulated miRNAs. Expression levels of selected miRNAs were validated by quantitative real-time PCR (qRT-PCR) in all serum and in articular cartilage samples from OA patients (n = 12) and healthy individuals (n = 7). Bioinformatics analysis was used to investigate the involved pathways and target genes for the above miRNAs. Results: We identified 279 differentially expressed miRNAs in the serum of OA patients compared to controls. Two hundred and five miRNAs (73.5%) were upregulated and 74 (26.5%) downregulated. ROC analysis revealed that 77 miRNAs had area under the curve (AUC) > 0.8 and p < 0.05. Bioinformatics analysis in the 77 miRNAs revealed that their target genes were involved in multiple signaling pathways associated with OA, among which FoxO, mTOR, Wnt, pI3K/akt, TGF-β signaling pathways, ECM-receptor interaction, and fatty acid biosynthesis. qRT-PCR validation in seven selected out of the 77 miRNAs revealed 3 significantly downregulated miRNAs (hsa-miR-33b-3p, hsa-miR-671-3p, and hsa-miR-140-3p) in the serum of OA patients, which were in silico predicted to be enriched in pathways involved in metabolic processes. Target-gene analysis of hsa-miR-140-3p, hsa-miR-33b-3p, and hsa-miR-671-3p revealed that InsR and IGFR1 were common targets of all three miRNAs, highlighting their involvement in regulation of metabolic processes that contribute to OA pathology. Hsa-miR-140-3p and hsa-miR-671-3p expression levels were consistently downregulated in articular cartilage of OA patients compared to healthy individuals. Conclusions: A serum miRNA signature was established for the first time using high density resolution miR-arrays in OA patients. We identified a three-miRNA signature, hsa-miR-140-3p, hsa-miR-671-3p, and hsa-miR-33b-3p, in the serum of OA patients, predicted to regulate metabolic processes, which could serve as a potential biomarker for the evaluation of OA risk and progression.Peer reviewedFinal Published versio

Measurements of neutrino oscillation in appearance and disappearance channels by the T2K experiment with 6.6 x 10(20) protons on target

111 pages, 45 figures, submitted to Physical Review D. Minor revisions to text following referee comments111 pages, 45 figures, submitted to Physical Review D. Minor revisions to text following referee comments111 pages, 45 figures, submitted to Physical Review D. Minor revisions to text following referee commentsWe thank the J-PARC staff for superb accelerator performance and the CERN NA61/SHINE Collaboration for providing valuable particle production data. We acknowledge the support of MEXT, Japan; NSERC, NRC, and CFI, Canada; CEA and CNRS/IN2P3, France; DFG, Germany; INFN, Italy; National Science Centre (NCN), Poland; RSF, RFBR and MES, Russia; MINECO and ERDF funds, Spain; SNSF and SER, Switzerland; STFC, UK; and the U. S. Deparment of Energy, USA. We also thank CERN for the UA1/NOMAD magnet, DESY for the HERA-B magnet mover system, NII for SINET4, the WestGrid and SciNet consortia in Compute Canada, GridPP, UK, and the Emerald High Performance Computing facility in the Centre for Innovation, UK. In addition, participation of individual researchers and institutions has been further supported by funds from ERC (FP7), EU; JSPS, Japan; Royal Society, UK; and DOE Early Career program, USA

Measurement of the electron neutrino charged-current interaction rate on water with the T2K ND280 pi(0) detector

10 pages, 6 figures, Submitted to PRDhttp://journals.aps.org/prd/abstract/10.1103/PhysRevD.91.112010© 2015 American Physical Society11 pages, 6 figures, as accepted to PRD11 pages, 6 figures, as accepted to PRD11 pages, 6 figures, as accepted to PR

Chronic Maternal Depression Is Associated with Reduced Weight Gain in Latino Infants from Birth to 2 Years of Age

BACKGROUND: Latino children are at increased risk for mirconutrient deficiencies and problems of overweight and obesity. Exposures in pregnancy and early postpartum may impact future growth trajectories. OBJECTIVES: To evaluate the relationship between prenatal and postnatal maternal depressive symptoms experienced in pregnancy and infant growth from birth to 2 years of age in a cohort of Latino infants. METHODS: We recruited pregnant Latina mothers at two San Francisco hospitals and followed their healthy infants to 24 months of age. At 6, 12 and 24 months of age, infants were weighed and measured. Maternal depressive symptoms were assessed prenatally and at 4-6 weeks postpartum. Women who had high depressive symptoms at both time periods were defined as having chronic depression. Logistic mixed models were applied to compare growth curves and risk for overweight and underweight based on exposure to maternal depression. RESULTS: We followed 181 infants to 24 months. At 12 and 24 months, respectively, 27.4% and 40.5% were overweight, and 5.6% and 2.2% were underweight. Exposure to chronic maternal depression was associated with underweight (OR = 12.12, 95%CI 1.86-78.78) and with reduced weight gain in the first 2 years of life (Coef = -0.48, 95% CI -0.94-0.01) compared with unexposed infants or infants exposed to episodic depression (depression at one time point). Exposure to chronic depression was also associated with reduced risk for overweight in the first 2 years of life (OR 0.28, 95%CI 0.03-0.92). CONCLUSIONS: Exposure to chronic maternal depression in the pre- and postnatal period was associated with reduced weight gain in the first two years of life and greater risk for failure to thrive, in comparison with unexposed infants or those exposed episodically. The infants of mothers with chronic depression may need additional nutritional monitoring and intervention

Laser-directed hierarchical assembly of liquid crystal defects and control of optical phase singularities

Topological defect lines are ubiquitous and important in a wide variety of fascinating phenomena and theories in many fields ranging from materials science to early-universe cosmology, and to engineering of laser beams. However, they are typically hard to control in a reliable manner. Here we describe facile erasable “optical drawing” of self-assembled defect clusters in liquid crystals. These quadrupolar defect clusters, stabilized by the medium's chirality and the tendency to form twisted configurations, are shaped into arbitrary two-dimensional patterns, including reconfigurable phase gratings capable of generating and controlling optical phase singularities in laser beams. Our findings bridge the studies of defects in condensed matter physics and optics and may enable applications in data storage, singular optics, displays, electro-optic devices, diffraction gratings, as well as in both optically- and electrically-addressed pixel-free spatial light modulators

Aptamer-based multiplexed proteomic technology for biomarker discovery

Interrogation of the human proteome in a highly multiplexed and efficient manner remains a coveted and challenging goal in biology. We present a new aptamer-based proteomic technology for biomarker discovery capable of simultaneously measuring thousands of proteins from small sample volumes (15 [mu]L of serum or plasma). Our current assay allows us to measure ~800 proteins with very low limits of detection (1 pM average), 7 logs of overall dynamic range, and 5% average coefficient of variation. This technology is enabled by a new generation of aptamers that contain chemically modified nucleotides, which greatly expand the physicochemical diversity of the large randomized nucleic acid libraries from which the aptamers are selected. Proteins in complex matrices such as plasma are measured with a process that transforms a signature of protein concentrations into a corresponding DNA aptamer concentration signature, which is then quantified with a DNA microarray. In essence, our assay takes advantage of the dual nature of aptamers as both folded binding entities with defined shapes and unique sequences recognizable by specific hybridization probes. To demonstrate the utility of our proteomics biomarker discovery technology, we applied it to a clinical study of chronic kidney disease (CKD). We identified two well known CKD biomarkers as well as an additional 58 potential CKD biomarkers. These results demonstrate the potential utility of our technology to discover unique protein signatures characteristic of various disease states. More generally, we describe a versatile and powerful tool that allows large-scale comparison of proteome profiles among discrete populations. This unbiased and highly multiplexed search engine will enable the discovery of novel biomarkers in a manner that is unencumbered by our incomplete knowledge of biology, thereby helping to advance the next generation of evidence-based medicine