scholarly journals Global identification of functional microRNA::mRNA interactions in Drosophila

2018 ◽  
Author(s):  
Hans-Hermann Wessels ◽  
Svetlana Lebedeva ◽  
Antje Hirsekorn ◽  
Neelanjan Mukherjee ◽  
Uwe Ohler
Keyword(s):  
Mirna Target ◽  
Signal To Noise Ratio ◽  
Absolute Quantification ◽  
Mediated Effects ◽  
Target Sites ◽  
Experimental Annotation ◽  
Global Identification ◽  
And Function

AbstractMicroRNAs (miRNAs) are key mediators of post-transcriptional gene expression silencing. Although Drosophila has been of critical importance for miRNA discovery, biogenesis and function, there has been no comprehensive experimental annotation of functional miRNA target sites. To close this gap, we generated the first in vivo map of miRNA::mRNA interactions in Drosophila melanogaster, making use of crosslinked nucleotides in Argonaute (AGO) crosslinking and immunoprecipitation (CLIP) experiments that enable an unambiguous assignment of miRNAs to AGO binding sites at much higher signal-to-noise ratio than computational predictions alone.Absolute quantification of cellular miRNA levels showed the miRNA pool in Drosophila cell lines to be more diverse than previously reported. Benchmarking two different CLIP approaches, we identified a similar predictive potential to unambiguously assign thousands of miRNA::mRNA pairs from AGO1 interaction data at unprecedented depth. Quantitative RNA-Seq and subcodon-resolution ribosomal footprinting data upon AGO1 depletion enabled the determination of miRNA-mediated effects on target expression and translation. We thus provide the first comprehensive resource of miRNA target sites as well as their quantitative functional impact in Drosophila.

scholarly journals HILIC-Enabled 13C Metabolomics Strategies: Comparing Quantitative Precision and Spectral Accuracy of QTOF High- and QQQ Low-Resolution Mass Spectrometry

Metabolites ◽  
2019 ◽  
Vol 9 (4) ◽  
pp. 63 ◽  
Author(s):  
André Feith ◽  
Attila Teleki ◽  
Michaela Graf ◽  
Lorenzo Favilli ◽  
Ralf Takors
Keyword(s):  
Mass Spectrometry ◽  
Isotope Dilution Mass Spectrometry ◽  
Absolute Quantification ◽  
Spectral Accuracy ◽  
Time Resolved ◽  
13C Tracer ◽  
Accurate Quantification ◽  
Tracer Experiments

Dynamic 13C-tracer-based flux analyses of in vivo reaction networks still require a continuous development of advanced quantification methods applying state-of-the-art mass spectrometry platforms. Utilizing alkaline HILIC chromatography, we adapt strategies for a systematic quantification study in non- and 13C-labeled multicomponent endogenous Corynebacterium glutamicum extracts by LC-QTOF high resolution (HRMS) and LC-QQQ tandem mass spectrometry (MS/MS). Without prior derivatization, a representative cross-section of 17 central carbon and anabolic key intermediates were analyzed with high selectivity and sensitivity under optimized ESI-MS settings. In column detection limits for the absolute quantification range were between 6.8–304.7 (QQQ) and 28.7–881.5 fmol (QTOF) with comparable linearities (3–5 orders of magnitude) and enhanced precision using QQQ-MRM detection. Tailor-made preparations of uniformly (U)13C-labeled cultivation extracts for isotope dilution mass spectrometry enabled the accurate quantification in complex sample matrices and extended linearities without effect on method parameters. Furthermore, evaluation of metabolite-specific m+1-to-m+0 ratios (ISR1:0) in non-labeled extracts exhibited sufficient methodical spectral accuracies with mean deviations of 3.89 ± 3.54% (QTOF) and 4.01 ± 3.01% (QQQ). Based on the excellent HILIC performance, conformity analysis of time-resolved isotopic enrichments in 13C-tracer experiments revealed sufficient spectral accuracy for QQQ-SIM detection. However, only QTOF-HRMS ensures determination of the full isotopologue space in complex matrices without mass interferences.

scholarly journals Gold nanorods based diffusion reflection measurements: current status and perspectives for clinical applications

Nanophotonics ◽  
2017 ◽  
Vol 6 (5) ◽  
pp. 1031-1042 ◽  
Author(s):  
Rinat Ankri ◽  
Dror Fixler
Keyword(s):  
Optical Imaging ◽  
Gold Nanorods ◽  
Diffusion Theory ◽  
Signal To Noise Ratio ◽  
High Specificity ◽  
Current Status ◽  
Tissue Properties ◽  
Dr Method ◽  
And Function

AbstractOptical imaging is a powerful tool for investigating the structure and function of tissues. Tissue optical imaging technologies are generally discussed under two broad regimes: microscopic and macroscopic, while the latter is widely investigated in the field of light-tissue interaction. Among the developed optical technologies for tissue investigation, the diffusion reflectance (DR) method is a simple and safe technology. However, this method suffers from low specificity and low signal-to-noise ratio, so the extraction of the tissue properties is not an easy task. In this review, we describe the use of gold nanorods (GNRs) in DR spectroscopy. The GNRs present unique optical properties which enhance the scattering and absorption properties of a tissue. The GNRs can be easily targeted toward abnormal sites in order to improve the DR signal and to distinguish between the healthy and the abnormal sites in the tissue, with high specificity. This article describes the use of the DR-GNRs method for the detection of cancer and atherosclerosis, from light transfer theory, through the extraction of the tissue properties using the diffusion theory and up to DR in vivo measurements.

Deuterium MR in Vivo Imaging of the Rat Eye Using 2H2O

1995 ◽  
Vol 36 (4-6) ◽  
pp. 552-555 ◽  
Author(s):  
T. Obata ◽  
H. Ikehira ◽  
F. Shishido ◽  
N. Fukuda ◽  
Y. Ueshima ◽  
...  
Keyword(s):  
Mr Imaging ◽  
Water Movement ◽  
Pulse Sequence ◽  
Signal To Noise Ratio ◽  
Spin Echo ◽  
Dynamic Change ◽  
Intraperitoneal Administration ◽  
Compartment Model

In vivo euterium MR imaging (2H MR) was investigated in rats after intraperitoneal administration of deuterated saline, and a dynamic study of the water movement in rat eyes was performed. Deuterium MR imaging was carried out by means of a gradient-echo (GRE) and a spin-echo (SE) pulse sequence. The rat eye was imaged in 2H MR more selectively by SE than by GRE, but a lower signal-to-noise ratio was obtained in 2H MR imaging using the SE sequence. The MR signal intensity of the rat eye was followed by a 3-compartment model, which enabled determination of the flow rate constant of the water in the eye (0.359/min). Deuterium MR imaging is useful to visualize the dynamic change of water in rat eyes using 2H MR at the same magnetic field (2 T) that can also be used for conventional MR imaging in humans.

scholarly journals Multisite Kinetic Modeling of13C Metabolic MR Using [1-13C]Pyruvate

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
Pedro A. Gómez Damián ◽  
Jonathan I. Sperl ◽  
Martin A. Janich ◽  
Oleksandr Khegai ◽  
Florian Wiesinger ◽  
...  
Keyword(s):  
Kinetic Modeling ◽  
Signal To Noise Ratio ◽  
Signal To Noise ◽  
Metabolic Conversion ◽  
Conversion Rates ◽  
Site Exchange ◽  
Noise Ratio ◽  
Parameter Values

Hyperpolarized13C imaging allows real-timein vivomeasurements of metabolite levels. Quantification of metabolite conversion between [1-13C]pyruvate and downstream metabolites [1-13C]alanine, [1-13C]lactate, and [13C]bicarbonate can be achieved through kinetic modeling. Since pyruvate interacts dynamically and simultaneously with its downstream metabolites, the purpose of this work is the determination of parameter values through a multisite, dynamic model involving possible biochemical pathways present in MR spectroscopy. Kinetic modeling parameters were determined by fitting the multisite model to time-domain dynamic metabolite data. The results for different pyruvate doses were compared with those of different two-site models to evaluate the hypothesis that for identical data the uncertainty of a model and the signal-to-noise ratio determine the sensitivity in detecting small physiological differences in the target metabolism. In comparison to the two-site exchange models, the multisite model yielded metabolic conversion rates with smaller bias and smaller standard deviation, as demonstrated in simulations with different signal-to-noise ratio. Pyruvate dose effects observed previously were confirmed and quantified through metabolic conversion rate values. Parameter interdependency allowed an accurate quantification and can therefore be useful for monitoring metabolic activity in different tissues.

scholarly journals The cellular microenvironment and cell adhesion: a role for O-glycosylation

2011 ◽  
Vol 39 (1) ◽  
pp. 378-382 ◽  
Author(s):  
Liping Zhang ◽  
Kelly G. Ten Hagen
Keyword(s):  
Cell Adhesion ◽  
Cancer Progression ◽  
Cellular Microenvironment ◽  
Developmental Defects ◽  
Mediated Effects ◽  
Cell Layers ◽  
And Function ◽  
Integrin Ligand

Glycosylation is one of the most abundant protein modifications in Nature, having roles in protein stability, secretion and function. Alterations in mucin-type O-glycosylation are responsible for a number of human diseases and developmental defects, as well as associated with certain types of cancer. However, the mechanistic role of this form of glycosylation in many of these instances is unclear. Here we describe how one glycosyltransferase responsible for initiating mucin-type O-glycosylation (PGANT3), specifically modulates integrin-mediated cell adhesion by influencing the secretion and localization of an integrin ligand. The integrin ligand Tiggrin, is normally O-glycosylated and localized to the basal matrix, where adhesion of two opposing cell layers takes place. In pgant3 mutants, Tiggrin is no longer O-glycosylated and fails to be properly secreted to the basal cell layer interface, resulting in disruption of proper cell adhesion. pgant3-mediated effects are dependent on the enzymatic activity of PGANT3 and cannot be rescued by another pgant family member, indicating a unique role for this glycosyltransferase. These results provide in vivo evidence for the role of O-glycosylation in the secretion of specific extracellular matrix proteins, which thereby influences the composition of the cellular ‘microenvironment’ and modulates cell adhesion events. The studies described in this review provide insight into the long-standing association between aberrant O-glycosylation and tumorigenesis, as changes in tumour environment and cell adhesion are hallmarks of cancer progression.

Conformation of Ribosomes from Escherichia Coli

1974 ◽  
Vol 32 ◽  
pp. 210-211
Author(s):  
M. Boublik ◽  
W. Hellmann ◽  
F. Jenkins
Keyword(s):  
Binding Sites ◽  
Ribosomal Proteins ◽  
Fluorescent Dyes ◽  
Protein Biosynthesis ◽  
Three Dimensional ◽  
Spatial Arrangement ◽  
Dimensional Structure ◽  
Complete Understanding ◽  
And Function

The present knowledge of the three-dimensional structure of ribosomes is far too limited to enable a complete understanding of the various roles which ribosomes play in protein biosynthesis. The spatial arrangement of proteins and ribonuclec acids in ribosomes can be analysed in many ways. Determination of binding sites for individual proteins on ribonuclec acid and locations of the mutual positions of proteins on the ribosome using labeling with fluorescent dyes, cross-linking reagents, neutron-diffraction or antibodies against ribosomal proteins seem to be most successful approaches. Structure and function of ribosomes can be correlated be depleting the complete ribosomes of some proteins to the functionally inactive core and by subsequent partial reconstitution in order to regain active ribosomal particles.

Determination of the Best Emulsion for the Microscopy of Crystals

1981 ◽  
Vol 39 ◽  
pp. 32-33
Author(s):  
David A. Grano ◽  
Kenneth H. Downing
Keyword(s):  
Spatial Frequency ◽  
Information Transfer ◽  
Signal To Noise Ratio ◽  
Experimental Situation ◽  
Photographic Emulsion ◽  
Modulation Transfer ◽  
Signal To Noise ◽  
Frequency Space ◽  
Noise Ratio

The retrieval of high-resolution information from images of biological crystals depends, in part, on the use of the correct photographic emulsion. We have been investigating the information transfer properties of twelve emulsions with a view toward 1) characterizing the emulsions by a few, measurable quantities, and 2) identifying the “best” emulsion of those we have studied for use in any given experimental situation. Because our interests lie in the examination of crystalline specimens, we've chosen to evaluate an emulsion's signal-to-noise ratio (SNR) as a function of spatial frequency and use this as our critereon for determining the best emulsion.The signal-to-noise ratio in frequency space depends on several factors. First, the signal depends on the speed of the emulsion and its modulation transfer function (MTF). By procedures outlined in, MTF's have been found for all the emulsions tested and can be fit by an analytic expression 1/(1+(S/S0)2). Figure 1 shows the experimental data and fitted curve for an emulsion with a better than average MTF. A single parameter, the spatial frequency at which the transfer falls to 50% (S0), characterizes this curve.

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