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An approach for calculating tunneling amplitudes from a nonlocalized initial state is presented. Generalizing the matching conditions and equations of motion to allow for complex momentum permits a description of tunneling in the presence of so-called classical motion. Possible applications of the method are presented.
A model of the mechanisms involved in transient (vulcanian-type) explosive eruptions has enabled us to place constraints on the velocities of blocks ejected during such events on Earth as well as on the excess pressure and concentration of gas in the vent prior to the onset of the explosion. This model, suitably modified, can predict the results of similar eruptions occurring under the differing Venusian environmental conditions. It is found that the much higher atmospheric pressure (approximately 100 bars at the mean planetary radius) dominates the form of the resulting deposit in two main ways: (1) by inhibiting the expansion out of the vent of overpressured gas, hence reducing ejecta velocity; and (2) by retarding the flight of ejected blocks via drag effects. Thus, it is expected that such deposits will typically extend to approximately 200 m on Venus as compared to several km for documented terrestrial deposits.
Highlights: • In hepatocytes, TRPM2 channels are mainly localised in intracellular organelles. • Oxidative stress increases the number of TRPM2 channels in the plasma membrane. • TRPM2 trafficking to the PM may contribute to positive Ca{sup 2+}-dependent feedback. Transient Receptor Potential Melastatin (TRPM) 2 is a non-selective Ca{sup 2+} permeable cation channel and a member of the Transient Receptor Potential (TRP) channel family. TRPM2 has unique gating properties; it is activated by intracellular ADP-ribose (ADPR), whereas Ca{sup 2+} plays a role of an important co-factor in channel activation, increasing TRPM2 sensitivity to ADPR. TRPM2 is highly expressed in rat and mouse hepatocytes, where it has been shown to contribute to oxidative stress-induced cell death and liver damage due to paracetamol-overdose. The mechanisms regulating the activity of TRPM2 channels in hepatocytes, however, are not well understood. In this paper, we investigate the localisation of TRPM2 protein in hepatocytes. The presented results demonstrate that in rat hepatocytes under normal conditions, most of the TRPM2 protein is localised intracellularly. This was determined by confocal microscopy using TRPM2-and plasma membrane (PM)-specific antibodies and immunofluorescence, and biotinylation studies followed by western blotting. Interestingly, in hepatocytes treated with either H{sub 2}O{sub 2} or paracetamol, the amount of TRPM2 co-localised with PM is significantly increased, compared to the untreated cells. It is concluded that trafficking of TRPM2 to the PM could potentially contribute to a positive feedback mechanism mediating Ca{sup 2+} overload in hepatocytes under conditions of oxidative stress.
Paramyxoviruses replicate in the cytoplasm with no obvious requirement to interact with the nucleus. Nevertheless, the W protein of the highly lethal bat-borne paramyxovirus Nipah virus (NiV) is known to undergo specific targeting to the nucleus, mediated by a single nuclear localisation signal (NLS) within the C-terminal domain. Here, we report for the first time that additional sites modulate nucleocytoplasmic localisation of W. We show that the N-terminal domain interacts with importin α1 and contributes to nuclear accumulation of W, indicative of a novel N-terminal NLS. We also find that W undergoes exportin-1 mediated nuclear export, dependent on a leucine at position 174. Together, these data enable significant revision of the generally accepted model of W trafficking, with implications for understanding of the mechanisms of NiV immune evasion. - Highlights: • A new model for Nipah virus W protein nucleocytoplasmic trafficking is proposed. • Nipah W protein is shown to undergo active nuclear export via exportin-1. • Nipah W nuclear import is mediated by multiple nuclear localisation signals.
The spatial compartmentalisation of biochemical signalling pathways is essential for cell function. Nesprins are a multi-isomeric family of proteins that have emerged as signalling scaffolds, herein, we investigate the localisation and function of novel nesprin-2 N-terminal variants. We show that these nesprin-2 variants display cell specific distribution and reside in both the cytoplasm and nucleus. Immunofluorescence microscopy revealed that nesprin-2 N-terminal variants colocalised with β-catenin at cell-cell junctions in U2OS cells. Calcium switch assays demonstrated that nesprin-2 and β-catenin are lost from cell-cell junctions in low calcium conditions whereas emerin localisation at the NE remained unaltered, furthermore, an N-terminal fragment of nesprin-2 was sufficient for cell-cell junction localisation and interacted with β-catenin. Disruption of these N-terminal nesprin-2 variants, using siRNA depletion resulted in loss of β-catenin from cell-cell junctions, nuclear accumulation of active β-catenin and augmented β-catenin transcriptional activity. Importantly, we show that U2OS cells lack nesprin-2 giant, suggesting that the N-terminal nesprin-2 variants regulate β-catenin signalling independently of the NE. Together, these data identify N-terminal nesprin-2 variants as novel regulators of β-catenin signalling that tether β-catenin to cell-cell contacts to inhibit β-catenin transcriptional activity. - Highlights: • N-terminal nesprin-2 variants display cell specific expression patterns. • N-terminal spectrin repeats of nesprin-2 interact with β-catenin. • N-terminal nesprin-2 variants scaffold β-catenin at cell-cell junctions.. • Nesprin-2 variants play multiple roles in β-catenin signalling.
The fundamental nature of turbulent density fluctuations in standard Wendelstein 7-X (W7-X) stellarator discharges is investigated experimentally via phase contrast imaging (PCI) in combination with gyrokinetic simulations with the code GENE. We find that density fluctuations are ion-temperature-gradient-driven and radially localised in the outer half of the plasma. It is shown that the line-integrated PCI measurements cover the right range of wavenumbers and a favourable toroidal and poloidal location to capture some of the strongest density fluctuations in W7-X. Due to the radial localisation of fluctuations, measured wavenumber–frequency spectra exhibit a dominant phase velocity, which can be related to the E × B rotation velocity at the radial position of a well in the neoclassical radial electric field. The match is robust against variations of heating power and line-integrated density, which is partly due to the localisation of fluctuations and partly due to effects of the radial gradient in the E × B velocity profile on the wavenumber–frequency spectrum. The latter effect is studied with a newly built synthetic PCI diagnostic and global gyrokinetic simulations with GENE-3D.
Long-chain acyl-CoA synthetases (LACS) play diverse and fundamentally important roles in lipid metabolism. While their functions have been well established in bacteria, yeast and plants, the mechanisms by which LACS isozymes regulate lipid metabolism in unicellular oil-producing microalgae, including the diatom Phaeodactylum tricornutum , remain largely unknown. In P. tricornutum , a family of five genes ( ptACSL1 – ptACSL5 ) encodes LACS activities. For this work, we generated single lacs knockout/knockdown mutants using multiplexed CRISPR/Cas9 method, and determined their substrate specificities towards different fatty acids (FAs) and subcellular localisations. ptACSL3 is localised in the mitochondria and its disruption led to compromised growth and reduced triacylglycerol (TAG) content when cells were bubbled with air. The ptACSL3 mutants showed altered FA profiles in two galactoglycerolipids and phosphatidylcholine (PC) with significantly reduced distribution of 16:0 and 16:1. ptACSL5 is localised in the peroxisome and its knockdown resulted in reduced growth rate and altered molecular species of PC and TAG, indicating a role in controlling the composition of acyl-CoAs for lipid synthesis. Our work demonstrates the potential of generating gene knockout mutants with the mutation of large fragment deletion using multiplexed CRISPR/Cas9 and provides insight into the functions of LACS isozymes in lipid metabolism in the oleaginous microalgae.
Cobalt is an essential element for life and plays a crucial role in supporting the drive to clean energy, due to its importance in rechargeable batteries. Co is often associated with Fe in the environment, but the fate of Co in Fe-rich biogeochemically-active environments is poorly understood. To address this, synchrotron-based scanning X-ray microscopy (SXM) was used investigate the behaviour of cobalt at the nanoscale in Co-Fe(III)-oxyhydroxides undergoing microbial reduction. SXM can assess spatial changes in metal speciation and organic compounds helping to elucidate the electron transfer processes occurring at the cell-mineral interface and inform on the fate of cobalt in redox horizons. G. sulfurreducens was used to reduce synthetic Co-ferrihydrite as an analogue of natural cobalt-iron-oxides. Magnetite [Fe(II)/Fe(III) 3 O 4 ] production was confirmed by powder X-ray diffraction (XRD), SXM and X-ray magnetic circular dichroism (XMCD) data, where best fits of the latter suggested Co-bearing magnetite. Macro-scale XAS techniques suggested Co(III) reduction occurred and complementary SXM at the nanoscale, coupled with imaging, found localised biogenic Co(III) reduction at the cell-mineral interface via direct contact with outer membrane cytochromes. No discernible localised changes in Fe speciation were detected in the reordered cobalt-iron-oxides that were formed and at the end point of the experiment only 11% Co and 1.5% Fe had been solubilised. The solid phase retention, alongside the highly localised and preferential cobalt bioreduction observed at the nanoscale is consistent with retention of Co in redox zones. This work improves our fundamental molecular-scale understanding of the fate of Co in complex environmental systems and supports the development of biogenic Co-doped magnetite for industrial applications from drug delivery systems to magnetic recording media.
In the vicinity of Europa, Galileo observed bursty Alfvén-cyclotron wave power at the gyrofrequencies of a number of species including K + , math formula, Na + , and Cl + , indicating the localised pickup of these species. Additional evidence for the presence of Chlorine was the occurrence of both left-hand (LH) and right-hand (RH) polarised transverse wave power near the Cl + gyrofrequency, thought to be due to the pickup of both Cl + and the easily formed Chlorine anion, Cl – . To test this hypothesis we use one-dimensional hybrid (kinetic ion, massless fluid electron) simulations for both positive and negative pickup ions and self-consistently reproduce the growth of both LH and RH Alfvén-cyclotron waves in agreement with linear theory. We show how the simultaneous generation of LH and RH waves can result in non-gyrotropic ion distributions and increased wave amplitudes, and how even trace quantities of negative pickup ions are able to generate an observable RH signal. Here, through comparing simulated and observed wave amplitudes, we are able to place the first constraints on the densities of Chlorine pickup ions in localised regions at Europa.
Atmospheric radioxenon levels are monitored worldwide by the Comprehensive Nuclear-Test-Ban Treaty Organisation (CTBTO) to detect emissions from nuclear explo-sions. Our paper examines the global network that has been set up to take routine air samples and to determine the atmospheric radioxenon concentrations. It is hypothesised that the monitoring system aims at the detection and localisation of radioxenon relea-ses. Both capabilities are dependent on background levels, explosive yields, leakage rates, transport time, choice of xenon isotope and number and locations of monitoring stations. And for each parameter, the global capabilities to detect and localise emissions from nuclear explosions are analysed. Recommendations for the future of the background sources and the development of the monitoring system are derived.
Hard core bosons in a large class of one or two dimensional flat band systems have an upper critical density, below which the ground states can be described completely. At the critical density, the ground states are Wigner crystals. If one adds a particle to the system at the critical density, the ground state and the low lying multi particle states of the system can be described as a Wigner crystal with an additional pair of particles. The energy band for the pair is separated from the rest of the multi-particle spectrum. The proofs use a Gerschgorin type of argument for block diagonally dominant matrices. In certain one-dimensional or tree-like structures one can show that the pair is localised, for example in the chequerboard chain. For this one-dimensional system with periodic boundary condition the energy band for the pair is flat, the pair is localised.
Auxin is a crucial phytohormone regulating multiple aspects of plant growth and differentiation, including regulation of vascular cambium activity, xylogenesis and its responsiveness towards gravitropic stress. Although the regulation of these biological processes greatly depends on auxin and regulators of the auxin signalling pathway, many of their specific functions remain unclear. Therefore, the present study aims to functionally characterise Eucalyptus grandis AUX/INDOLE-3-ACETIC ACID 13 (EgrIAA13), a member of the auxin signalling pathway. In Eucalyptus and Populus, EgrIAA13 and its orthologs are preferentially expressed in the xylogenic tissues and downregulated in tension wood. Therefore, to further investigate EgrIAA13 and its function during xylogenesis, we conducted subcellular localisation and Induced Somatic Sector Analysis experiments using overexpression and RNAi knockdown constructs of EgrIAA13 to create transgenic tissue sectors on growing stems of Eucalyptus and Populus. Since Aux/IAAs interact with Auxin Responsive Factors (ARFs), in silico predictions of IAA13-ARF interactions were explored and experimentally validated via yeast-2-hybrid experiments. Our results demonstrate that EgrIAA13 localises to the nucleus and that downregulation of EgrIAA13 impedes Eucalyptus xylem fibre and vessel development. We also observed that EgrIAA13 interacts with Eucalyptus ARF2, ARF5, ARF6 and ARF19A. Based on these results, we conclude that EgrIAA13 is a regulator of Eucalyptus xylogenesis and postulate that the observed phenotypes are likely to result from alterations in the auxin-responsive transcriptome via IAA13-ARF modules such as EgrIAA13-EgrARF5. Our results provide the first insights into the regulatory role of EgrIAA13 during xylogenesis.
Pin1-type parvulins are unique among PPIases that can catalyse an otherwise slow cis-trans isomerisation of phosphorylated peptide bond preceding proline in target proteins. This prolyl isomerisation process can regulate activity, stability and localisation of target proteins and thus control cellular processes like eukaryotic cell proliferation, cell cycle progression and gene regulation. Towards understanding the function of Pin1-type prolyl isomerisation in Dictyostelium discoideum, a slime mould with distinct growth and developmental phases, we identified PinA as a novel Pin1-type parvulin by its ability to complement the temperature sensitivity phenotype associated with a mutation in ESS1 in S. cerevisiae. In D. discoideum, pinA is temporally and spatially regulated during growth and development. PinA is both nuclear as well as cytoplasmic in the growing cells. We further show that loss of pinA (pinA{sup −}) leads to decreased growth rate, reduced spore formation and abnormal prespore-prestalk patterning. We conclude that PinA is required for normal growth as well as development in D. discoideum. - Highlights: • PinA is a bona fide homologue of S. cerevisiae Ess1. • PinA is required for normal cell proliferation of D. discoideum. • PinA is spatially localised in developmental structures. • PinA is important for cell differentiation and patterning.
Understanding the impact of repository gas, generated from degradation of waste and its interaction with the host rock, is essential when assessing the performance and safety function of long-term disposal systems for radioactive waste. Numerical models based on conventional multi-phase flow theory have historically been applied to predict the outcome and impact of gas flow on different repository components. However, they remain unable to describe the full complexity of the physical processes observed in water-saturated experiments (e.g., creation of dilatant pathways) and thus, the development of novel representations for their description is required when assessing fully saturated clay-based systems. This was the primary focus of Task A within the international cooperative project DECOVALEX-2019 (D-2019) and refinement of these approaches is the primary focus of this study (Task B in the current phase of DECOVALEX-2023). This paper summarises development of enhanced numerical representations of key processes and compares the performance of each model against high-quality laboratory test data. Experimental data reveals that gas percolation in water-saturated compacted bentonite is characterised by four key features: (i) a quiescence phase, followed by (ii) the gas breakthrough, which leads to a (iii) peak value, which is then followed by (iv) a negative decay. Three models based on the multiphase flow theory have been developed. These models can provide good initial values and reasonable responses for gas breakthrough (although some of them still predict a too-smooth response). Peak gas pressure values are in general reasonably well captured, although maximum radial stress differences are observed at 48 mm from the base of the sample. Here, numerical peak values of 12.8 MPa are predicted, whereas experimental values are about 11 MPa. These models are also capable of providing a reasonable representation of the negative pressure decay following peak pressure. However, other key specific features (such as the timing of gas breakthrough) still require a better representation. The model simulations and their comparison with experimental data show that these models need to be further improved with respect to model parameter calibration, the numerical representation of spatial heterogeneities in material properties and flow localisation, and the upscaling of the related physical processes and parameters. To further understand gas flow localisation, a new conceptual model has been developed, which shows that discrete channels can possibly be induced through the instability of gas-bentonite interface during gas injection, thus providing a new perspective for modeling gas percolation in low-permeability deformable media.
Highlights: • Recovery processes in precipitate free zones differ from those in grain interiors. • Most precipitate free zones contain few dislocations, after deformation. • Deformation can induce large misorientations in precipitate free zones. • Some deformed precipitate free zones contain nanoscale grains. - Abstract: Scanning precession electron diffraction and automated crystal orientation mapping in a transmission electron microscope (TEM) have been applied to quantitatively study the crystal orientation of precipitate free zones (PFZs) of four GB regions in an AA6060 alloy in peak aged condition (temper T6) after uniaxial compression to 20% engineering strain. The PFZ width in the alloy was found to be w = 170 ± 40 nm. The results show that some PFZs develop significant misorientations relative to their parent grain, and represent, to the best knowledge of the authors, the first quantitative evidence of this. This misorientation may either be constant inside a particular PFZ, making it appear like a band or a very elongated subgrain, or be partitioned in discrete regions with a diameter comparable to the PFZ width, making the former PFZ into a collection of small grains. The band-like PFZ observed in this work had a misorientation relative to its parent grain of ≈ 7°, while the grain-like PFZ had grains with misorientations between ≈ 12° and ≈ 20° relative to its parent grain. The other PFZs that were observed had only limited misorientations relative to their parent grains, and had either dislocations perpendicular to the GB plane or a dislocation wall at the transition region. A general TEM study of the material at various engineering strains was also conducted and suggests that grain-like PFZs are more frequent for larger strains, indicating that the different PFZ features are likely due to different strain localisation in individual PFZs. This localisation is expected to be influenced by the orientation of the loading axis relative to crystal orientations and GB planes. It is also suggested that the different PFZ features engender different work hardening rates and possibly affect nucleation of intergranular fracture. The results support previous studies on the microstructure evolution of PFZs in age-hardenable aluminium alloys during deformation.
Highlights: • Brief oestrogen treatment causes the cytoplasmic retention of the testis factor SOX9. • The cytoplasmic retention of SOX9 by oestrogen is associated with the rapid stabilisation of microtubules. • ERK1/2 is required for the stabilisation of microtubules by activated oestrogen receptors. • Oestrogen can act on both a non-genomic and genomic level to impact gonad somatic cell fate. Nuclear SOX9 is essential for Sertoli cell differentiation and the development of a testis. Exposure of Sertoli cells to exogenous oestrogen causes cytoplasmic retention of SOX9, inhibiting testis development and promoting ovarian development. The cytoplasmic localisation of SOX9 requires a stabilised microtubule network and a key MAPK complex, ERK1/2, is responsive to oestrogen and known to affect the microtubule network. We hypothesised that oestrogen could stabilise microtubules through the activation of ERK1/2 to promote the cytoplasmic retention of SOX9. Treatment of human testis-derived NT2/D1 cells for 30 min with oestrogen rapidly activated ERK1/2, stabilised the microtubule network and increased cytoplasmic localisation of SOX9. The effects of oestrogen on SOX9 and tubulin were blocked by the ERK1/2 inhibitor U0126, demonstrating that ERK1/2 mediates the stabilisation of microtubules and cytoplasmic retention of SOX9 by oestrogen. Together, these data revealed a previously unknown mechanism for oestrogen in impacting MAPK signalling to block SOX9 bioavailability and the differentiation of Sertoli cells.
Highlights: • Rab30 localised primarily to TGN and recycling endosomes. • Rab30 plays a major role in the spatial organization of the TGN and recycling endosomes. • Deficiency of Rab30 results in a perturbation in endosome-TGN retrograde transport. • Interactive partners of Rab30 include Arf1 and Arf4. • Findings highlight the interconnected network between the TGN and endosomal system. Rab30 is a poorly characterized small GTPase. Here we show that Rab30 is localised primarily to the TGN and recycling endosomes in a range of cell types, including primary neurons; minor levels of Rab30 were also detected throughout the Golgi stack and early endosomes. Silencing of Rab30 resulted in the dispersal of both early and recycling endosomes and TGN compartments in HeLa cells. By analyzing cargo trafficking in Rab30-silenced and Rab30-overexpressing HeLa cells, we demonstrate that Rab30 plays a role in retrograde trafficking of TGN38 from endosomes to the Golgi, but has no apparent role in the endocytic recycling of the transferrin receptor to the plasma membrane. Five interactive partners with Rab30 were identified by pull-down and MS analysis using GFP-tagged Rab30 mutant, Rab30(Q68L). Two of the interactive partners identified were Arf1 and Arf4, known regulators of endosome to TGN retrograde transport. Knockdown of Arf1 and Arf4 results in GFP-Rab30 decorated tubules arising from the recycling endosomes, suggesting association of Rab30 with tubular carriers. Overall our data demonstrates a role for Rab30 in regulating retrograde transport to the TGN and maintenance of endosomal-TGN organization.
Curvature-driven instabilities are ubiquitous in magnetised fusion plasmas. By analysing the conservation laws of the gyrokinetic system of equations, we demonstrate that the well-known spatial localisation of these instabilities to regions of ‘bad magnetic curvature’ can be explained using the conservation law for a sign-indefinite quadratic quantity that we call the gyrokinetic field invariant. Its evolution equation allows us to define the local effective magnetic curvature whose sign demarcates the regions of ‘good’ and ‘bad’ curvature, which, under some additional simplifying assumptions, can be shown to correspond to the inboard (high-field) and outboard (low-field) sides of a tokamak plasma, respectively. We find that, given some reasonable assumptions, electrostatic curvature-driven modes are always localised to the regions of bad magnetic curvature, regardless of the specific character of the instability. More importantly, we also deduce that any mode that is unstable in the region of good magnetic curvature must be electromagnetic in nature. As a concrete example, we present the magnetic-drift mode, a novel good-curvature electromagnetic instability, and compare its properties with the well-known electron-temperature-gradient instability. Finally, we discuss the relevance of the magnetic drift mode for high-β fusion plasmas, and in particular its relationship with microtearing modes.