The many structural faces of calmodulin: a multitasking molecular jackknife

Calmodulin (CaM) is a highly conserved protein and a crucial calcium sensor in eukaryotes. CaM is a regulator of hundreds of diverse target proteins. A wealth of studies has been carried out on the structure of CaM, both in the unliganded form and in complexes with target proteins and peptides. The outcome of these studies points toward a high propensity to attain various conformational states, depending on the binding partner. The purpose of this review is to provide examples of different conformations of CaM trapped in the crystal state. In addition, comparisons are made to corresponding studies in solution. The different CaM conformations in crystal structures are also compared based on the positions of the metal ions bound to their EF hands, in terms of distances, angles, and pseudo-torsion angles. Possible caveats and artifacts in CaM crystal structures are discussed, as well as the possibilities of trapping biologically relevant CaM conformations in the crystal state.     

Phase transitions and antiferromagnetism in copper(II) hexanoates: a new tetranuclear type of copper carboxylate paddle-wheel association

A series of compounds of formula [{Cu₂(OOCC_mH_2m + 1)₄(urea)}₂] (m=5-11) have been characterized. X-ray structure analysis for the hexanoate compound reveals a new type of tetranuclear dicopper(II) tetracarboxylate, where the central coordination sphere in [{Cu₂(OOCC₅H₁₁)₄(urea)}₂] is composed of two dinuclear dicopper tetracarboxylates, connected via two inter-dinuclear Cu-O coordination bonds at a distance 2.222(2) Å through the apical positions of two dimers. Urea molecules (Cu-O 2.114(2) Å) occupy both outside apical positions of the resulting tetranuclear units. A strong antiferromagnetic behaviour has been shown for [{Cu₂(OOCC₅H₁₁)₄(urea)}₂] (−2J=261.4(4) cm⁻¹), and compared with related isolated dinuclear and polymeric hexanoate compounds [Cu₂(OOCC₅H₁₁)₄(urea)₂], [Cu₂(OOCC₅H₁₁)₄]_n, respectively. Only small differences in the magnetic susceptibility have been found, while EPR spectroscopy showed significantly different results for all three hexanoate compounds, also with the dicopper tetracarboxylate central core and square-pyramidal CuO₄O chromophores. A solid-to-solid phase transition for [{Cu₂(OOCC₅H₁₁)₄(urea)}₂] was observed by magnetic measurement and analysed for the whole series [{Cu₂(OOCC_mH_2m + 1)₄(urea)}₂] by TG, DTA, and variable temperature XRD studies.     

Lumbo-sacral neural crest derivatives fate mapped with the aid of Wnt-1 promoter integrate but are not essential to kidney development

Neural crest (NC) cells may be involved in kidney organogenesis by providing inductive signals and contributing to cells of the renal stroma. We show here that the lumbo-sacral NC cells fate mapped with the aid of Wnt-1 promoter in the mouse migrate close to the metanephros at the initiation of organogenesis but these cells remain superficial to the condensed Pax2-expressing mesenchymal cells. NC-derived cells enter later into the kidney proper from the midline region. The NC cells contribute also to development of the extra-adrenal para-aortic bodies, Zuckerkandl's bodies and the nerve cord of the sympathetic nervous system. Splotch (Sp^2H/Sp^2H) embryos, having a NC defect in the lumbo-sacral region, develop a normal metanephros even though the kidney does not express the NC markers Sox10, Phox2b and tyrosine hydroxylase. Consistent with the histological findings, the kidneys of Sp^2H/Sp^2H embryos also express the stromal genes Foxd1, Hoxa10 and RARβ normally. Wnt-1 promoter-marked wild-type LacZ NC cells migrate intensely from the heterologous inducer tissue of the embryonic dorsal spinal cord (SPC) to the kidney mesenchyme, but tubule induction does not depend on NC migration, since the Sp^2H/Sp^2H SPC also induces tubulogenesis. The Sp^2H/Sp^2H mesenchyme also remains competent for tubulogenesis. We conclude that the NC cells fate mapped with the aid of Wnt-1 promoter migrate to the close to the metanephros and form later derivatives integrating with the kidney, but they may not be essential to the development of the stromal cells nor they may provide critical morphogenetic signals to regulate early kidney development in vivo.     

Linkage disequilibrium network analysis (LDna) gives a global view of chromosomal inversions,local adaptation and geographic structure

Recent advances in sequencing allow population‐genomic data to be generated for virtually any species. However, approaches to analyse such data lag behind the ability to generate it, particularly in nonmodel species. Linkage disequilibrium (LD, the nonrandom association of alleles from different loci) is a highly sensitive indicator of many evolutionary phenomena including chromosomal inversions, local adaptation and geographical structure. Here, we present linkage disequilibrium network analysis (LDna), which accesses information on LD shared between multiple loci genomewide. In LD networks, vertices represent loci, and connections between vertices represent the LD between them. We analysed such networks in two test cases: a new restriction‐site‐associated DNA sequence (RAD‐seq) data set for Anopheles baimaii, a Southeast Asian malaria vector; and a well‐characterized single nucleotide polymorphism (SNP) data set from 21 three‐spined stickleback individuals. In each case, we readily identified five distinct LD network clusters (single‐outlier clusters, SOCs), each comprising many loci connected by high LD. In A. baimaii, further population‐genetic analyses supported the inference that each SOC corresponds to a large inversion, consistent with previous cytological studies. For sticklebacks, we inferred that each SOC was associated with a distinct evolutionary phenomenon: two chromosomal inversions, local adaptation, population‐demographic history and geographic structure. LDna is thus a useful exploratory tool, able to give a global overview of LD associated with diverse evolutionary phenomena and identify loci potentially involved. LDna does not require a linkage map or reference genome, so it is applicable to any population‐genomic data set, making it especially valuable for nonmodel species.     

Genetic analysis of sea-ice bacterial communities of the Western Baltic Sea using an improved double gradient method

The bacterial diversity of sea ice from Kiel Bight obtained during the rare event of solid ice cover in spring 1996 was analysed by molecular genetic approaches using an improved double gradient denaturing gradient gel electrophoretic method (DG-DGGE) to separate 16S rDNA fragments of approximately 500 bp. The excellent separation of individual bands within these gradient gels allowed us to obtain sequence information and to allocate the phylogenetic position of representative bacteria from the sea ice. The band pattern of the gradient gels revealed a vertical stratification of the bacterial species distribution within the ice and the presence of characteristic bacteria for each layer. According to their 16S rDNA sequences, major bands of the gradient gels represented bacteria closely related to fermenting species of the genera Propionibacterium and Bacteroides and to anoxygenic phototrophic purple sulfur bacteria (Chromatiaceae). Their abundance in horizons of the inner ice core may indicate the existence of oxygen-deficient and anoxic zones or niches and possible primary production by anoxygenic photosynthesis within the investigated Baltic Sea sea ice. This is the first phylogenetic evidence of the presence, and most probably the development, of phototrophic purple sulfur bacteria in sea ice.     

Role of the V-ATPase in regulation of the vacuolar fission-fusion equilibrium

Like numerous other eukaryotic organelles, the vacuole of the yeast Saccharomyces cerevisiae undergoes coordinated cycles of membrane fission and fusion in the course of the cell cycle and in adaptation to environmental conditions. Organelle fission and fusion processes must be balanced to ensure organelle integrity. Coordination of vacuole fission and fusion depends on the interactions of vacuolar SNARE proteins and the dynamin-like GTPase Vps1p. Here, we identify a novel factor that impinges on the fusion-fission equilibrium: the vacuolar H(+)-ATPase (V-ATPase) performs two distinct roles in vacuole fission and fusion. Fusion requires the physical presence of the membrane sector of the vacuolar H(+)-ATPase sector, but not its pump activity. Vacuole fission, in contrast, depends on proton translocation by the V-ATPase. Eliminating proton pumping by the V-ATPase either pharmacologically or by conditional or constitutive V-ATPase mutations blocked salt-induced vacuole fragmentation in vivo. In living cells, fission defects are epistatic to fusion defects. Therefore, mutants lacking the V-ATPase display large single vacuoles instead of multiple smaller vacuoles, the phenotype that is generally seen in mutants having defects only in vacuolar fusion. Its dual involvement in vacuole fission and fusion suggests the V-ATPase as a potential regulator of vacuolar morphology and membrane dynamics.     

Isolation,patch size and matrix effects on bird assemblages in forest reserves

Habitat fragmentation is one of the most studied topics in ecology but our knowledge is still limited particularly concerning matrix effects on species distribution in a human-dominated landscape. We tested the ability of random sampling hypothesis, colonization–extinction dynamics and matrix-related concepts to explain the variation in species richness, total bird density and community composition in old-forest bird assemblages in two contrasting landscapes. We collected data on breeding bird abundances from 66 old-growth forest reserves in NE Finland and six larger areas in adjacent Russian Karelia using the line transect method. In Finland, protected old-forest patches are embedded in a matrix dominated by young regeneration stands. In Russia, study areas were situated in continuous, old forest dominated landscapes. Bird assemblages in old-forest patches embedded in human-modified matrix in Finland were not random samples from Russian bird assemblages. In the Finnish assemblages, species richness was lower and total bird density higher. Species richness declined with increasing distance (isolation) to Russia. Bird assemblages in large forest reserves in Finland close to Russia were structurally more similar to assemblages in the continuous reference landscape than those in small and more distant reserves. The results support the idea that several mechanisms related to colonisation–extinction dynamics and to matrix resource availability influence species distribution in fragmented landscapes but in species-specific ways. We conclude that even though small and isolated protected areas may foster high relative bird species density their ecological integrity is compromised, and therefore, improving matrix quality around reserves may lead to considerable conservation benefits.     

Linkage disequilibrium clustering‐based approach for association mapping with tightly linked genomewide data

Genomewide association studies (GWAS) aim to identify genetic markers strongly associated with quantitative traits by utilizing linkage disequilibrium (LD) between candidate genes and markers. However, because of LD between nearby genetic markers, the standard GWAS approaches typically detect a number of correlated SNPs covering long genomic regions, making corrections for multiple testing overly conservative. Additionally, the high dimensionality of modern GWAS data poses considerable challenges for GWAS procedures such as permutation tests, which are computationally intensive. We propose a cluster‐based GWAS approach that first divides the genome into many large nonoverlapping windows and uses linkage disequilibrium network analysis in combination with principal component (PC) analysis as dimensional reduction tools to summarize the SNP data to independent PCs within clusters of loci connected by high LD. We then introduce single‐ and multilocus models that can efficiently conduct the association tests on such high‐dimensional data. The methods can be adapted to different model structures and used to analyse samples collected from the wild or from biparental F₂ populations, which are commonly used in ecological genetics mapping studies. We demonstrate the performance of our approaches with two publicly available data sets from a plant (Arabidopsis thaliana) and a fish (Pungitius pungitius), as well as with simulated data.