114 research outputs found
Harnessing nuclear spin polarization fluctuations in a semiconductor nanowire
Soon after the first measurements of nuclear magnetic resonance (NMR) in a
condensed matter system, Bloch predicted the presence of statistical
fluctuations proportional to in the polarization of an ensemble of
spins. First observed by Sleator et al., so-called "spin noise" has
recently emerged as a critical ingredient in nanometer-scale magnetic resonance
imaging (nanoMRI). This prominence is a direct result of MRI resolution
improving to better than 100 nm^3, a size-scale in which statistical spin
fluctuations begin to dominate the polarization dynamics. We demonstrate a
technique that creates spin order in nanometer-scale ensembles of nuclear spins
by harnessing these fluctuations to produce polarizations both larger and
narrower than the natural thermal distribution. We focus on ensembles
containing ~10^6 phosphorus and hydrogen spins associated with single InP and
GaP nanowires (NWs) and their hydrogen-containing adsorbate layers. We monitor,
control, and capture fluctuations in the ensemble's spin polarization in
real-time and store them for extended periods. This selective capture of large
polarization fluctuations may provide a route for enhancing the weak magnetic
signals produced by nanometer-scale volumes of nuclear spins. The scheme may
also prove useful for initializing the nuclear hyperfine field of electron spin
qubits in the solid-state.Comment: 18 pages, 5 figure
Universal Vectorial and Ultrasensitive Nanomechanical Force Field Sensor
Miniaturization of force probes into nanomechanical oscillators enables
ultrasensitive investigations of forces on dimensions smaller than their
characteristic length scale. Meanwhile it also unravels the force field
vectorial character and how its topology impacts the measurement. Here we
expose an ultrasensitive method to image 2D vectorial force fields by
optomechanically following the bidimensional Brownian motion of a singly
clamped nanowire. This novel approach relies on angular and spectral tomography
of its quasi frequency-degenerated transverse mechanical polarizations:
immersing the nanoresonator in a vectorial force field does not only shift its
eigenfrequencies but also rotate eigenmodes orientation as a nano-compass. This
universal method is employed to map a tunable electrostatic force field whose
spatial gradients can even take precedence over the intrinsic nanowire
properties. Enabling vectorial force fields imaging with demonstrated
sensitivities of attonewton variations over the nanoprobe Brownian trajectory
will have strong impact on scientific exploration at the nanoscale
Nanomechanical sensing using spins in diamond
Nanomechanical sensors and quantum nanosensors are two rapidly developing
technologies that have diverse interdisciplinary applications in biological and
chemical analysis and microscopy. For example, nanomechanical sensors based
upon nanoelectromechanical systems (NEMS) have demonstrated chip-scale mass
spectrometry capable of detecting single macromolecules, such as proteins.
Quantum nanosensors based upon electron spins of negatively-charged
nitrogen-vacancy (NV) centers in diamond have demonstrated diverse modes of
nanometrology, including single molecule magnetic resonance spectroscopy. Here,
we report the first step towards combining these two complementary technologies
in the form of diamond nanomechanical structures containing NV centers. We
establish the principles for nanomechanical sensing using such
nano-spin-mechanical sensors (NSMS) and assess their potential for mass
spectrometry and force microscopy. We predict that NSMS are able to provide
unprecedented AC force images of cellular biomechanics and to, not only detect
the mass of a single macromolecule, but also image its distribution. When
combined with the other nanometrology modes of the NV center, NSMS potentially
offer unparalleled analytical power at the nanoscale.Comment: Errors in the stress susceptibility parameters present in the
original arXiv version have been correcte
Histone Deacetylase Inhibitors Downregulate Checkpoint Kinase 1 Expression to Induce Cell Death in Non-Small Cell Lung Cancer Cells
Background: Histone deacetylase inhibitors (HDACis) are promising anticancer drugs; however, the molecular mechanisms leading to HDACi-induced cell death have not been well understood and no clear mechanism of resistance has been elucidated to explain limited efficacy of HDACis in clinical trials. Methods and Findings: Here, we show that protein levels of checkpoint kinase 1 (Chk1), which has a major role in G2 cell cycle checkpoint regulation, was markedly reduced at the protein and transcriptional levels in lung cancer cells treated with pan-and selective HDACis LBH589, scriptaid, valproic acid, apicidin, and MS-275. In HDACi treated cells Chk1 function was impaired as determined by decreased inhibitory phosphorylation of cdc25c and its downstream target cdc2 and increased expression of cdc25A and phosphorylated histone H3, a marker of mitotic entry. In time course experiments, Chk1 downregulation occurred after HDACi treatment, preceding apoptosis. Ectopic expression of Chk1 overcame HDACiinduced cell death, and pretreating cells with the cdc2 inhibitor purvalanol A blocked entry into mitosis and prevented cell death by HDACis. Finally, pharmacological inhibition of Chk1 showed strong synergistic effect with LBH589 in lung cancer cells. Conclusions: These results define a pathway through which Chk1 inhibition can mediate HDACi-induced mitotic entry and cell death and suggest that Chk1 could be an early pharmacodynamic marker to assess HDACi efficacy in clinical samples
Targeting Sphingosine Kinase 1 in Carcinoma Cells Decreases Proliferation and Survival by Compromising PKC Activity and Cytokinesis
Sphingosine kinases (SK) catalyze the phosphorylation of proapoptotic sphingosine to the prosurvival factor sphingosine 1-phosphate (S1P), thereby promoting oncogenic processes. Breast (MDA-MB-231), lung (NCI-H358), and colon (HCT 116) carcinoma cells were transduced with shRNA to downregulate SK-1 expression or treated with a pharmacologic SK-1 inhibitor. The effects of SK-1 targeting were investigated by measuring the level of intracellular sphingosine, the activity of protein kinase C (PKC) and cell cycle regulators, and the mitotic index. Functional assays included measurement of cell proliferation, colony formation, apoptosis, and cell cycle analysis. Downregulation of SK-1 or its pharmacologic inhibition increased intracellular sphingosine and decreased PKC activity as shown by reduced phosphorylation of PKC substrates. In MDA-MB-231 cells this effect was most pronounced and reduced cell proliferation and colony formation, which could be mimicked using exogenous sphingosine or the PKC inhibitor RO 31-8220. SK-1 downregulation in MDA-MB-231 cells increased the number of cells with 4N and 8N DNA content, and similar effects were observed upon treatment with sphingosine or inhibitors of SK-1 or PKC. Examination of cell cycle regulators unveiled decreased cdc2 activity and expression of Chk1, which may compromise spindle checkpoint function and cytokinesis. Indeed, SK-1 kd cells entered mitosis but failed to divide, and in the presence of taxol also failed to sustain mitotic arrest, resulting in further increased endoreduplication and apoptosis. Our findings delineate an intriguing link between SK-1, PKC and components of the cell cycle machinery, which underlines the significance of SK-1 as a target for cancer therapy
Array-based molecular karyotyping in fetuses with isolated brain malformations identifies disease-causing CNVs
ZO-1 controls endothelial adherens junctions, cell–cell tension, angiogenesis, and barrier formation
Identification of a Lacosamide Binding Protein Using an Affinity Bait and Chemical Reporter Strategy: 14-3-3 ζ
We have advanced a useful strategy to elucidate binding partners of ligands (drugs) with modest binding affinity. Key to this strategy is attaching to the ligand an affinity bait (AB) and a chemical reporter (CR) group, where the AB irreversibly attaches the ligand to the receptor upon binding and the CR group is employed for receptor detection and isolation. We have tested this AB&CR strategy using lacosamide ((R)-1), a low-molecular-weight antiepileptic drug. We demonstrate that using a (R)-lacosamide AB&CR agent ((R)-2) 14-3-3 ζ in rodent brain soluble lysates is preferentially adducted, adduction is stereospecific with respect to the AB&CR agent, and adduction depends upon the presence of endogenous levels of the small molecule metabolite xanthine. Substitution of lacosamide AB agent ((R)- 5) for (R)-2 led to the identification of the 14-3-3 ζ adduction site (K120) by mass spectrometry. Competition experiments using increasing amounts of (R)-1 in the presence of (R)-2 demonstrated that (R)-1 binds at or near the (R)-2 modification site on 14-3-3 ζ. Structure-activity studies of xanthine derivatives provided information concerning the likely binding interaction between this metabolite and recombinant 14-3-3 ζ. Documentation of the 14-3-3 ζ-xanthine interaction was obtained with isothermal calorimetry using xanthine and the xanthine analogue 1,7-dimethylxanthine
Simvastatin Sodium Salt and Fluvastatin Interact with Human Gap Junction Gamma-3 Protein
Finding pleiomorphic targets for drugs allows new indications or warnings for treatment to be identified. As test of concept, we applied a new chemical genomics approach to uncover additional targets for the widely prescribed lipid-lowering pro-drug simvastatin. We used mRNA extracted from internal mammary artery from patients undergoing coronary artery surgery to prepare a viral cardiovascular protein library, using T7 bacteriophage. We then studied interactions of clones of the bacteriophage, each expressing a different cardiovascular polypeptide, with surface-bound simvastatin in 96-well plates. To maximise likelihood of identifying meaningful interactions between simvastatin and vascular peptides, we used a validated photo-immobilisation method to apply a series of different chemical linkers to bind simvastatin so as to present multiple orientations of its constituent components to potential targets. Three rounds of biopanning identified consistent interaction with the clone expressing part of the gene GJC3, which maps to Homo sapiens chromosome 7, and codes for gap junction gamma-3 protein, also known as connexin 30.2/31.3 (mouse connexin Cx29). Further analysis indicated the binding site to be for the N-terminal domain putatively ‘regulating’ connexin hemichannel and gap junction pores. Using immunohistochemistry we found connexin 30.2/31.3 to be present in samples of artery similar to those used to prepare the bacteriophage library. Surface plasmon resonance revealed that a 25 amino acid synthetic peptide representing the discovered N-terminus did not interact with simvastatin lactone, but did bind to the hydrolysed HMG CoA inhibitor, simvastatin acid. This interaction was also seen for fluvastatin. The gap junction blockers carbenoxolone and flufenamic acid also interacted with the same peptide providing insight into potential site of binding. These findings raise key questions about the functional significance of GJC3 transcripts in the vasculature and other tissues, and this connexin’s role in therapeutic and adverse effects of statins in a range of disease states
Tight junctions: from simple barriers to multifunctional molecular gates
Epithelia and endothelia separate different tissue compartments and protect multicellular organisms from the outside world. This requires the formation of tight junctions, selective gates that control paracellular diffusion of ions and solutes. Tight junctions also form the border between the apical and basolateral plasma-membrane domains and are linked to the machinery that controls apicobasal polarization. Additionally, signalling networks that guide diverse cell behaviours and functions are connected to tight junctions, transmitting information to and from the cytoskeleton, nucleus and different cell adhesion complexes. Recent advances have broadened our understanding of the molecular architecture and cellular functions of tight junctions
- …