Direct Single-Molecule Observation of Mode and Geometry of RecA-Mediated Homology Search

Genomic integrity, when compromised by accrued DNA lesions, is maintained through efficient repair via homologous recombination. For this process the ubiquitous recombinase A (RecA), and its homologues such as the human Rad51, are of central importance, able to align and exchange homologous sequences within single-stranded and double-stranded DNA in order to swap out defective regions. Here, we directly observe the widely debated mechanism of RecA homology searching at a single-molecule level using high-speed atomic force microscopy (HS-AFM) in combination with tailored DNA origami frames to present the reaction targets in a way suitable for AFM-imaging. We show that RecA nucleoprotein filaments move along DNA substrates via short-distance facilitated diffusions, or slides, interspersed with longer-distance random moves, or hops. Importantly, from the specific interaction geometry, we find that the double-stranded substrate DNA resides in the secondary DNA binding-site within the RecA nucleoprotein filament helical groove during the homology search. This work demonstrates that tailored DNA origami, in conjunction with HS-AFM, can be employed to reveal directly conformational and geometrical information on dynamic protein-DNA interactions which was previously inaccessible at an individual single-molecule level

Cooperative RecA clustering: the key to efficient homology searching

The mechanism by which pre-synaptic RecA nucleoprotein filaments efficiently locate sequence homology across genomic DNA remains unclear. Here, using atomic force microscopy, we directly investigate the intermediates of the RecA-mediated homologous recombination process and find it to be highly cooperative, involving multiple phases. Initially, the process is dominated by a rapid 'association' phase, where multiple filaments interact on the same dsDNA simultaneously. This cooperative nature is reconciled by the observation of localized dense clusters of pre-synaptic filaments interacting with the observed dsDNA molecules. This confinement of reactive species within the vicinity of the dsDNA, is likely to play an important role in ensuring that a high interaction rate between the nucleoprotein filaments and the dsDNA can be achieved. This is followed by a slower 'resolution' phase, where the synaptic joints either locate sequence homology and progress to a post-synaptic joint, or dissociate from the dsDNA. Surprisingly, the number of simultaneous synaptic joints decreases rapidly after saturation of the dsDNA population, suggesting a reduction in interaction activity of the RecA filaments. We find that the time-scale of this decay is in line with the time-scale of the dispersion of the RecA filament clusters, further emphasising the important role this cooperative phenomena may play in the RecA-facilitated homology search

European mineral statistics 2009-13 : a product of the World Mineral Statistics database

This volume is the latest edition of a series that began in 2002 following the replacement of ‘World Mineral Statistics’ with ‘World Mineral Production’. It contains mineral production, import and export data for more than 70 mineral commodities, for 36 European countries including all EU Member States and EU Candidate Countries, plus Norway and Switzerland. These data are presented in two sections: by individual country and by commodity; the latter is illustrated by graphics. It remains the only freely available and up-to-date publication dedicated to statistical information about minerals and metals in Europe

European Mineral Statistics 2010-14: a product of the World Mineral statistics database

European Mineral Statistics provides statistical information about minerals and metals in Europe. It provides the essential background intelligence for any European minerals-related activities. Production, export and import tables are presented for all EU members and EU candidate countries, plus Norway and Switzerland, in two sections: •by individual country •by commodity, with bullets on salient features and graphics More than 70 different mineral commodities are included from Aluminium to Zirconium, plus statistics relating to primary aggregates and cement. The book also includes commentary discussing the different categories of minerals – construction minerals, industrial minerals, metals and energy minerals – in the European context and contains general information on the compilation of data

Mineral resource information in support of national, regional and local planning : Cheshire (comprising Cheshire, Boroughs of Halton and Warrington)

This report is one of a series prepared by the British Geological Survey for various administrative areas in England for the Office of the Deputy Prime Minister’s research project Mineral Resource Information in Support of National, Regional and Local Planning. The accompanying map relates to the county of Cheshire (comprising Cheshire, Boroughs of Halton and Warrington), and delineates the mineral resources of current, or potential, economic interest in the area and the sites where minerals are or have been worked. It also relates these to national planning designations, which may represent constraints on the extraction of minerals. Three major elements of information are presented: • the geological distribution and importance of mineral resources; • the extent of mineral planning permissions and the location of current mineral workings; and • the extent of selected, nationally-designated planning constraints. This wide range of information, much of which is scattered and not always available in a consistent and convenient form, is presented on a digitally-generated summary map on the scale of 1:100 000. This scale is convenient for the overall display of the data and allows for a legible topographic base on which to depict the information. However, all the data are held digitally at larger scales using a Geographical Information System (GIS), which allows easy revision, updating and customisation of the information together with its possible integration with other datasets. The information will form part of a Summary of the Mineral Resources of the North West Region. The purpose of the work is to assist all interested parties involved in the preparation and review of development plans, both in relation to the extraction of minerals and the protection of mineral resources from sterilisation. It provides a knowledge base, in a consistent format, on the nature and extent of mineral resources and the environmental constraints, which may affect their extraction. An important objective is to provide baseline data for the long term. The results may also provide a starting point for discussion on specific planning proposals for minerals extraction or on proposals, which may sterilise resources. It is anticipated that the map and report will also provide valuable background data for a much wider audience, including the different sectors of the minerals industry, other agencies and authorities (e.g. The Planning Inspectorate Agency, the Environment Agency, The Countryside Agency and English Nature), environmental interests and the general public. Basic mineral resource information is essential to support mineral exploration and development activities, for resource management and land-use planning, and to establish baseline data for environmental impact studies and environmental guidelines. It also enables a more sustainable pattern and standard of development to be achieved by valuing mineral resources as national assets. The mineral resources covered are sand and gravel, crushed rock aggregate, silica sand, salt, brick clay, building stones, peat, coal, hydrocarbons and metalliferous mineralisation

Mineral resource information in support of national, regional and local planning : Devon (comprising Devon, Plymouth, Torbay, Dartmoor National Park and part of Exmoor National Park)

This report is one of a series prepared by the British Geological Survey for various administrative areas in England for the Office of the Deputy Prime Minister’s research project Mineral Resource Information in Support of National, Regional and Local Planning. The accompanying maps relate to the county of Devon and delineates the mineral resources of current, or potential, economic interest in the area and the sites where minerals are or have been worked. It also relates these to national planning designations, which may represent constraints on the extraction of minerals. Three major elements of information are presented: • the geological distribution and importance of mineral resources; • the extent of mineral planning permissions and the location of current mineral workings; and • the extent of selected, nationally-designated planning constraints. This wide range of information, much of which is scattered and not always available in a consistent and convenient form, is presented on two digitally-generated summary map on the scale of 1:100 000. This scale is convenient for the overall display of the data and allows for a legible topographic base on which to depict the information. However, all the data are held digitally at larger scales using a Geographical Information System (GIS), which allows easy revision, updating and customisation of the information together with its possible integration with other datasets. The information will form part of a Summary of the Mineral Resources of the South West Region. The purpose of the work is to assist all interested parties involved in the preparation and review of development plans, both in relation to the extraction of minerals and the protection of mineral resources from sterilisation. It provides a knowledge base, in a consistent format, on the nature and extent of mineral resources and the environmental constraints, which may affect their extraction. An important objective is to provide baseline data for the long term. The results may also provide a starting point for discussion on specific planning proposals for minerals extraction or on proposals, which may sterilise resources. It is anticipated that the map and report will also provide valuable background data for a much wider audience, including the different sectors of the minerals industry, other agencies and authorities (e.g. The Planning Inspectorate Agency, the Environment Agency, The Countryside Agency and English Nature), environmental interests and the general public. Basic mineral resource information is essential to support mineral exploration and development activities, for resource management and land-use planning, and to establish baseline data for environmental impact studies and environmental guidelines. It also enables a more sustainable pattern and standard of development to be achieved by valuing mineral resources as national assets. The mineral resources covered are sand and gravel, crushed rock aggregate, brick clay, kaolin, ball clay, building stones, hydrocarbons and metalliferous mineralisation

From Monochrome to Technicolor: Simple Generic Approaches to Multicomponent Protein Nanopatterning Using Siloxanes with Photoremovable Protein-Resistant Protecting Groups.

We show that sequential protein deposition is possible by photodeprotection of films formed from a tetraethylene-glycol functionalized nitrophenylethoxycarbonyl-protected aminopropyltriethoxysilane (NPEOC-APTES). Exposure to near-UV irradiation removes the protein-resistant protecting group, and allows protein adsorption onto the resulting aminated surface. The protein resistance was tested using proteins with fluorescent labels and microspectroscopy of two-component structures formed by micro- and nanopatterning and deposition of yellow and green fluorescent proteins (YFP/GFP). Nonspecific adsorption onto regions where the protecting group remained intact was negligible. Multiple component patterns were also formed by near-field methods. Because reading and writing can be decoupled in a near-field microscope, it is possible to carry out sequential patterning steps at a single location involving different proteins. Up to four different proteins were formed into geometric patterns using near-field lithography. Interferometric lithography facilitates the organization of proteins over square cm areas. Two-component patterns consisting of 150 nm streptavidin dots formed within an orthogonal grid of bars of GFP at a period of ca. 500 nm could just be resolved by fluorescence microscopy

Influence of enhanced melt supply on upper crustal structure at a mid-ocean ridge discontinuity: A three-dimensional seismic tomographic study of 9°N East Pacific Rise

We present a three-dimensional upper crustal model of the 9°03′N overlapping spreading center (OSC) on the East Pacific Rise that assists in understanding the relationship between melt sills and upper crustal structure at a ridge discontinuity with enhanced melt supply at crustal levels. Our P wave velocity model obtained from tomographic inversion of ∼70,000 crustal first arrival travel times suggests that the geometry of extrusive emplacement are significantly different beneath the overlapping spreading limbs. Extrusive volcanic rocks above the western melt sill are inferred to be thin (∼250 m). More extensive accumulation of extrusives is inferred to the west than to the east of the western melt sill. The extrusive layer inferred above the eastern melt sill thickens from ∼350 (at the neovolcanic axis) to 550 m (to the west of the melt sill). Volcanic construction is likely to be significant in the formation of ridge crest morphology at the OSC, particularly at the tip of the eastern limb. On the basis of our interpretation of the velocity model, we propose that enhanced magma supply at crustal levels at the OSC may provide an effective mechanism for the migration of ridge discontinuities. This “dynamic magma supply model” may explain the commonly observed nonsteady migration pattern of ridge discontinuities by attributing this to the temporal fluctuations in melt availability to the overlapping spreading limbs

Formation of the Stomatal Outer Cuticular Ledge Requires a Guard Cell Wall Proline-Rich Protein

Stomata are formed by a pair of guard cells which have thickened, elastic cell walls to withstand the large increases in turgor pressure that have to be generated to open the pore that they surround. We have characterised FOCL1, a guard cell-expressed, secreted protein with homology to hydroxyproline-rich cell wall proteins. FOCL1-GFP localises to the guard cell outer cuticular ledge and plants lacking FOCL1 produce stomata without a cuticular ledge. Instead the majority of stomatal pores are entirely covered-over by a continuous fusion of the cuticle, and consequently plants have decreased levels of transpiration and display drought tolerance. The focl1 guard cells are larger and less able to reduce the aperture of their stomatal pore in response to closure signals suggesting that the flexibility of guard cell walls is impaired. FOCL1 is also expressed in lateral root initials where it aids lateral root emergence. We propose that FOCL1 acts in these highly specialised cells of the stomata and root to impart cell wall strength at high turgor and/or to facilitate interactions between the cell wall and the cuticle

Pattern formation in directional solidification under shear flow. I: Linear stability analysis and basic patterns

An asymptotic interface equation for directional solidification near the absolute stabiliy limit is extended by a nonlocal term describing a shear flow parallel to the interface. In the long-wave limit considered, the flow acts destabilizing on a planar interface. Moreover, linear stability analysis suggests that the morphology diagram is modified by the flow near the onset of the Mullins-Sekerka instability. Via numerical analysis, the bifurcation structure of the system is shown to change. Besides the known hexagonal cells, structures consisting of stripes arise. Due to its symmetry-breaking properties, the flow term induces a lateral drift of the whole pattern, once the instability has become active. The drift velocity is measured numerically and described analytically in the framework of a linear analysis. At large flow strength, the linear description breaks down, which is accompanied by a transition to flow-dominated morphologies, described in a companion paper. Small and intermediate flows lead to increased order in the lattice structure of the pattern, facilitating the elimination of defects. Locally oscillating structures appear closer to the instability threshold with flow than without.Comment: 20 pages, Latex, accepted for Physical Review