Flexibility within the Rotor and Stators of the Vacuolar H+-ATPase

The V-ATPase is a membrane-bound protein complex which pumps protons across the membrane to generate a large proton motive force through the coupling of an ATP-driven 3-stroke rotary motor (V₁) to a multistroke proton pump (V_o). This is done with near 100% efficiency, which is achieved in part by flexibility within the central rotor axle and stator connections, allowing the system to flex to minimise the free energy loss of conformational changes during catalysis. We have used electron microscopy to reveal distinctive bending along the V-ATPase complex, leading to angular displacement of the V₁ domain relative to the V_o domain to a maximum of ~30°. This has been complemented by elastic network normal mode analysis that shows both flexing and twisting with the compliance being located in the rotor axle, stator filaments, or both. This study provides direct evidence of flexibility within the V-ATPase and by implication in related rotary ATPases, a feature predicted to be important for regulation and their high energetic efficiencies.     

Functional Genomic Analysis of the let-7 Regulatory Network in Caenorhabditis elegans

The let-7 microRNA (miRNA) regulates cellular differentiation across many animal species. Loss of let-7 activity causes abnormal development in Caenorhabditis elegans and unchecked cellular proliferation in human cells, which contributes to tumorigenesis. These defects are due to improper expression of protein-coding genes normally under let-7 regulation. While some direct targets of let-7 have been identified, the genome-wide effect of let-7 insufficiency in a developing animal has not been fully investigated. Here we report the results of molecular and genetic assays aimed at determining the global network of genes regulated by let-7 in C. elegans. By screening for mis-regulated genes that also contribute to let-7 mutant phenotypes, we derived a list of physiologically relevant potential targets of let-7 regulation. Twenty new suppressors of the rupturing vulva or extra seam cell division phenotypes characteristic of let-7 mutants emerged. Three of these genes, opt-2, prmt-1, and T27D12.1, were found to associate with Argonaute in a let-7–dependent manner and are likely novel direct targets of this miRNA. Overall, a complex network of genes with various activities is subject to let-7 regulation to coordinate developmental timing across tissues during worm development.     

Quantifying Policy Options for Reducing Future Coronary Heart Disease Mortality in England: A Modelling Study

Aims

To estimate the number of coronary heart disease (CHD) deaths potentially preventable in England in 2020 comparing four risk factor change scenarios.

Methods and Results

Using 2007 as baseline, the IMPACT_SEC model was extended to estimate the potential number of CHD deaths preventable in England in 2020 by age, gender and Index of Multiple Deprivation 2007 quintiles given four risk factor change scenarios: (a) assuming recent trends will continue; (b) assuming optimal but feasible levels already achieved elsewhere; (c) an intermediate point, halfway between current and optimal levels; and (d) assuming plateauing or worsening levels, the worst case scenario. These four scenarios were compared to the baseline scenario with both risk factors and CHD mortality rates remaining at 2007 levels. This would result in approximately 97,000 CHD deaths in 2020. Assuming recent trends will continue would avert approximately 22,640 deaths (95% uncertainty interval: 20,390-24,980). There would be some 39,720 (37,120-41,900) fewer deaths in 2020 with optimal risk factor levels and 22,330 fewer (19,850-24,300) in the intermediate scenario. In the worst case scenario, 16,170 additional deaths (13,880-18,420) would occur. If optimal risk factor levels were achieved, the gap in CHD rates between the most and least deprived areas would halve with falls in systolic blood pressure, physical inactivity and total cholesterol providing the largest contributions to mortality gains.

Conclusions

CHD mortality reductions of up to 45%, accompanied by significant reductions in area deprivation mortality disparities, would be possible by implementing optimal preventive policies.     

Development of a multi-assay approach for monitoring coral diversity using eDNA metabarcoding

Coral Reefs - Cumulative anthropogenic pressures have triggered a global decline in the health of marine ecosystems, and coral reefs, in particular, are in crisis. With climate and...     

Medial epithelial seam cell migration during palatal fusion

The mammalian secondary palate forms from two shelves of mesenchyme sheathed in a single-layered epithelium. These shelves meet during embryogenesis to form the midline epithelial seam (MES). Failure of MES degradation prevents mesenchymal confluence and results in a cleft palate. Previous studies indicated that MES cells undergo features of epithelial-to-mesenchymal transition (EMT) and may become migratory as part of the fusion mechanism. To detect MES cell movement over the course of fusion, we imaged the midline of fusing embryonic ephrin-B2/GFP mouse palates in real time using two-photon microscopy. These mice express an ephrin-B2-driven green fluorescent protein (GFP) that labels the palatal epithelium nuclei and persists in those cells through the time window necessary for fusion. We observed collective migration of MES cells toward the oral surface of the palatal shelf over 48 hr of imaging, and we confirmed histologically that the imaged palates had fused by the end of the imaged period. We previously reported that ephrin reverse signaling in the MES is required for palatal fusion. We therefore added recombinant EphA4/Fc protein to block this signaling in imaged palates. The blockage inhibited fusion, as expected, but did not change the observed migration of GFP-labeled cells. Thus, we uncoupled migration and fusion. Our data reveal that palatal MES cells undergo a collective, unidirectional movement during palatal fusion and that ephrin reverse signaling, though required for fusion, controls aspects of the fusion mechanism independent of migration.     

Survival and behavior of juvenile steelhead trout (Oncorhynchus mykiss) in two estuaries in Oregon, USA

Anadromous salmonids are viewed as a prized commodity and cultural symbol throughout the Pacific coast of North America. Unfortunately, several native salmonid populations are threatened or at risk of extinction. Despite this, little is known about the behavior and survival of these fish as the juveniles transition from freshwater to the ocean. Our primary objectives were to estimate survival of juvenile steelhead migrating between trapping sites and the ocean and evaluate whether survival in the estuary varies temporally (within a year) or spatially (within and between estuaries) within the same distinct population segment. We also evaluated whether flow or fork length were correlated with survival and collected information on variables that have been demonstrated to affect smolt survival in other studies to lend insight regarding differences in survival estimates between basins. We compared run timing, migration rate, survival, condition factor, age composition and time of residence in the estuary for steelhead outmigrants from each basin and measured parasite loads in outmigrating steelhead to evaluate potential differences in parasite density and parasite community between basins. In 2009, we implanted acoustic transmitters in 139 wild steelhead smolts in two small rivers on the Oregon Coast. In general, only 40–50 % of the wild steelhead smolts tagged at upstream smolt traps were detected entering the ocean. The majority of mortality occurred in the lower estuary near the ocean. Wild steelhead smolts typically spent less than 1 day in the estuary in both basins. Using similar data from previous studies in the Nehalem and Alsea basins, we showed that survival appears to be negatively correlated with flow in most releases, and in 2009 fork length was not correlated with survival. Our observations provide baseline information on factors that could influence smolt survival through the estuary as well as smolt to adult survival in these basins, and emphasize the importance of monitoring smolt survival in the estuary.     

Molecular dynamics simulation on the effect of transition metal binding to the N-terminal fragment of amyloid-β

Abstract

We report molecular dynamics simulations of three possible adducts of Fe(II) to the N-terminal 1–16 fragments of the amyloid-β peptide, along with analogous simulations of Cu(II) and Zn(II) adducts. We find that multiple simulations from different starting points reach pseudo-equilibration within 100–300?ns, leading to over 900?ns of equilibrated trajectory data for each system. The specifics of the coordination modes for Fe(II) have only a weak effect on peptide secondary and tertiary structures, and we therefore compare one of these with analogous models of Cu(II) and Zn(II) complexes. All share broadly similar structural features, with mixture of coil, turn and bend in the N-terminal region and helical structure for residues 11–16. Within this overall pattern, subtle effects due to changes in metal are evident: Fe(II) complexes are more compact and are more likely to occupy bridge and ribbon regions of Ramachandran maps, while Cu(II) coordination leads to greater occupancy of the poly-proline region. Analysis of representative clusters in terms of molecular mechanics energy and atoms-in-molecules properties indicates similarity of four-coordinate Cu and Zn complexes, compared to five-coordinate Fe complex that exhibits lower stability and weaker metal–ligand bonding.

Communicated by Ramaswamy H. Sarma     

A Chemokine-Like Viral Protein Enhances Alpha Interferon Production by Plasmacytoid Dendritic Cells but Delays CD8+ T Cell Activation and Impairs Viral Clearance

Murine cytomegalovirus encodes numerous proteins that act on a variety of pathways to modulate the innate and adaptive immune responses. Here, we demonstrate that a chemokine-like protein encoded by murine cytomegalovirus activates the early innate immune response and delays adaptive immunity, thereby impairing viral clearance. The protein, m131/129 (also known as MCK-2), is not required to establish infection in the spleen; however, a mutant virus lacking m131/129 was cleared more rapidly from this organ. In the absence of m131/129 expression, there was enhanced activation of dendritic cells (DC), and virus-specific CD8⁺ T cells were recruited into the immune response earlier. Viral mutants lacking m131/129 elicited weaker production of alpha interferon (IFN-α) at 40 h postinfection, indicating that this protein exerts its effects during early rounds of viral replication in the spleen. Furthermore, while wild-type and mutant viruses activated plasmacytoid dendritic cells (pDC) equally at this time, as measured by the upregulation of costimulatory molecules, the presence of m131/129 stimulated more pDC to secrete IFN-α, accounting for the stronger IFN-α response than from the wild-type virus. These data provide evidence for a novel immunomodulatory function of a viral chemokine and expose the multifunctionality of immune evasion proteins. In addition, these results broaden our understanding of the interplay between innate and adaptive immunity.