DNA sequence variation in BpMADS2 gene in two populations of Betula pendula

The PISTILLATA (PI) homologue, BpMADS2, was isolated from silver birch (Betula pendula Roth) and used to study nucleotide polymorphism. Two regions (together about 2450 bp) comprising mainly untranslated sequences were sequenced from 10 individuals from each of two populations in Finland. The nucleotide polymorphism was low in the BpMADS2 locus, especially in the coding region. The synonymous site overall nucleotide diversity (pis) was 0.0043 and the nonsynonymous nucleotide diversity (pia) was only 0.000052. For the whole region, the pi values for the two populations were 0.0039 and 0.0045, and for the coding regions, the pi values were only 0 and 0.00066 (for the corresponding coding regions of Arabidopsis thaliana PI world-wide pi was 0.0021). Estimates of pi or theta did not differ significantly between the two populations, and the two populations were not diverged from each other. Two classes of BpMADS2 alleles were present in both populations, suggesting that this gene exhibits allelic dimorphism. In addition to the nucleotide site variation, two microsatellites were also associated within the haplotypes. This allelic dimorphism might be the result of postglacial re-colonization partly from northwestern, partly from southeastern/eastern refugia. The sequence comparison detected five recombination events in the regions studied. The large number of microsatellites in all of the three introns studied suggests that BpMADS2 is a hotspot for microsatellite formation.     

Expression of normal and mutant GFP-tagged y(+)L amino acid transporter-1 in mammalian cells

Lysinuric protein intolerance (LPI; MIM 222700) is an autosomal recessive disorder characterized by defective transport of cationic amino acids lysine, arginine and ornithine. The defect is localized in the basolateral membrane of polar epithelial cells of the renal tubules and intestine. The SLC7A7 (solute carrier family 7, member 7) gene that encodes y(+)LAT-1 (y(+)L amino acid transporter-1) is mutated in LPI, and leads to the malfunction of the heterodimer composed of y(+)LAT-1 and 4F2hc (4F2 heavy chain) responsible for the system y(+)L amino acid transport activity at the membrane. In this study, the intracellular trafficking and membrane expression of wild type and four mutant y(+)LAT-1 proteins (LPI(Fin), G54V, 1548delC, W242X) was studied in two human cell lines by expressing green fluorescent protein (GFP) tagged proteins. Different SLC7A7 mutations influenced the trafficking of y(+)LAT-1 in the cells differently, as the wild type and missense mutant fusion proteins localized to the plasma membrane, while the frameshift and nonsense mutants sequestered to the cytoplasmic membranes, never reaching the target areas of the cell.     

Severe inbreeding depression in collared flycatchers (Ficedula albicollis)

The causes and magnitude of inbreeding depression are of considerable importance for a wide range of issues in evolutionary and conservation biology, but we have only a limited understanding of inbreeding depression in natural populations. Here, we present a study of inbreeding in a large wild population of collared flycatchers (Ficedula albicollis). Inbreeding was rare, to the extent that we detected only 1.04% of 2139 matings over 18 years that resulted in offspring with a non-zero inbreeding coefficient, f > 0. When it did occur, inbreeding caused a significant reduction in the egg-hatching rate, in fledgling skeletal size and in post-fledging juvenile survival, with the number of offspring being recruited to the breeding population from a nest of f = 0.25 being reduced by 94% relative to a non-inbred nest. A maximum-likelihood estimate of the number of lethal equivalents per gamete was very high at B = 7.47, indicating a substantial genetic load in this population. There was also a non-significant tendency for inbreeding depression to increase with the strength of selection on a trait. The probability of mating between close relatives (f = 0.25) increased throughout the breeding season, possibly reflecting increased costs of inbreeding avoidance. Our results illustrate how severe inbreeding depression and considerable genetic load may exist in natural populations, but detecting them may require extensive long-term datasets.     

The influence of landscape structure on occurrence, abundance and genetic diversity of the common frog, Rana temporaria

Human‐caused habitat destruction and modification constitute one of the largest threats to population persistence and biodiversity, and are also suspected to be the major cause behind the global decline of amphibian populations. We assessed the potential role of agriculture‐related habitat fragmentation on population size and genetic variability in the common frog (Rana temporaria) by recording the occurrence, population density and genetic diversity in three geographically disparate regions in Sweden – each containing landscapes of high and low agricultural activity – and related these to landscape variables extracted from digital maps. We found a highly significant region‐by‐landscape interaction in occurrence, population density and genetic diversity revealing a reversed response to agriculture from south to north: while the effects of agriculture on R. temporaria populations were negative in the south, there were no effects in the central region, whereas positive effects were observed in the north. Spatial autocorrelation analyses of genetic data revealed that populations in high agricultural activity areas were more isolated than populations in low activity areas both in the southern and central regions of Sweden. Landscape diversity showed a strong positive correlation with both density and occurrence of frogs in Sweden as a whole, as well as in the southern region. Also, negative effects of roads and positive effects of ditches on genetic diversity were found in the south. Overall, these results suggest clear but regionally opposite effects of habitat structure on the population size and genetic diversity of amphibian populations. This means that the management strategy aiming to maximize the size and genetic diversity of local common frog populations, and perhaps also those of other amphibian populations, should account for regional differences in existing land‐use patterns.     

Influence of DPH on the structure and dynamics of a DPPC bilayer

We have conducted extensive molecular dynamics (MD) simulations together with differential scanning calorimetry (DSC) and nuclear magnetic resonance (NMR) experiments to quantify the influence of free 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescent probes on the structure and dynamics of a dipalmitoylphosphatidylcholine bilayer. Atomistic MD simulations show that in the membrane-water interface the influence of DPH is minor, whereas in the acyl-chain region DPH gives rise to major perturbations. In the latter case, DPH is found to influence a wide range of membrane properties, such as the packing and ordering of hydrocarbon tails and the lateral diffusion of lipid molecules. The effects are prominent but of local nature, i.e., the changes observed in the properties of lipid molecules are significant in the vicinity of DPH, but reduce rapidly as the distance from the probe increases. Long-range perturbations due to DPH are hence not expected. Detailed DSC and (2)H NMR measurements support this view. DSC shows only subtle perturbation to the cooperative behavior of the membrane system in the presence of DPH, and (2)H NMR shows that DPH gives rise to a slight increase in the lipid chain order, in agreement with MD simulations. Potential effects of other probes such as pyrene are briefly discussed.     

Intermolecular interactions of lysobisphosphatidic acid with phosphatidylcholine in mixed bilayers

Lysobisphosphatidic acid (LBPA) can be regarded to represent a unique derivative of phosphatidylglycerol. This lipid is highly enriched in late endosomes where it can comprise up to 10-15 mol% of all lipids and in these membranes, LBPA appears to be segregated into microdomains. We studied the thermotropic behavior of pure dioleoyl-LBPA mono- and bilayers using Langmuir-lipid monolayers, electron microscopy, differential scanning calorimetry (DSC), and fluorescence spectroscopy. LBPA formed metastable, liquid-expanded monolayers at an air/buffer interface, and its compression isotherms lacked any indication for structural phase transitions. Neat LBPA formed multilamellar vesicles with no structural transitions or phase transitions between 10 and 80 degrees C at a pH range of 3.0-7.4. We then proceeded to study mixed LBPA/dipalmitoylphosphatidylcholine (DPPC) bilayers by DSC and fluorescence spectroscopy. Incorporating increasing amounts of LBPA (up to X(LBPA) (molar fraction)=0.10) decreased the co-operativity of the main transition for DPPC, and a decrease in the main phase transition as well as pretransition temperature of DPPC was observed yet with no effect on the enthalpy of this transition. In keeping with the DSC data for DPPC, 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC)/LBPA mixed bilayers were more fluid, and no evidence for lateral phase segregation was observed. These results were confirmed using fluorescence microscopy of Langmuir-lipid films composed of POPC and LBPA up to X(LBPA)=0.50 with no evidence for lateral phase separation. As late endosomes are eminently acidic, we examined the effect of lowering pH on lateral organization of mixed PC/LBPA bilayers by DSC and fluorescence spectroscopy. Even at pH 3.0, we find no evidence of LBPA-induced microdomain formation at LBPA contents found in cellular organelles.     

Polymorphisms in the tyrosine kinase 2 and interferon regulatory factor 5 genes are associated with systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a complex systemic autoimmune disease caused by both genetic and environmental factors. Genome scans in families with SLE point to multiple potential chromosomal regions that harbor SLE susceptibility genes, and association studies in different populations have suggested several susceptibility alleles for SLE. Increased production of type I interferon (IFN) and expression of IFN-inducible genes is commonly observed in SLE and may be pivotal in the molecular pathogenesis of the disease. We analyzed 44 single-nucleotide polymorphisms (SNPs) in 13 genes from the type I IFN pathway in 679 Swedish, Finnish, and Icelandic patients with SLE, in 798 unaffected family members, and in 438 unrelated control individuals for joint linkage and association with SLE. In two of the genes—the tyrosine kinase 2 (TYK2) and IFN regulatory factor 5 (IRF5) genes—we identified SNPs that displayed strong signals in joint analysis of linkage and association (unadjusted P<10^-7) with SLE. TYK2 binds to the type I IFN receptor complex and IRF5 is a regulator of type I IFN gene expression. Thus, our results support a disease mechanism in SLE that involves key components of the type I IFN system.     

Habitat utilisation by fish community in a short-term regulated river reservoir

Short-term streamflow regulation (hydropeaking) affects the ecology of regulated rivers. We examined the longitudinal and temporal changes occurring in fish assemblages in a hydropeaking single river reservoir between two power plants by using electrofishing along the shoreline, hydroacoustics and test fishing in the open water. A longitudinally changing fish community was found among bottom-dwelling fish in the fast-flowing and highly disturbed upstream part of the reservoir progressing to generalists and pelagic fish in the lentic and most stable environment at the downstream end. The fish assemblage showed temporal patterns as fish density increased during night-time darkness and also towards autumn. Our work provides evidence for gradient effects of flow regulation and contributes to awareness of the effects of disturbance (flow/habitat variability) on biological systems.     

Dissecting the EphA3/Ephrin-A5 interactions using a novel functional mutagenesis screen

The EphA3 receptor tyrosine kinase preferentially binds ephrin-A5, a member of the corresponding subfamily of membrane-associated ligands. Their interaction regulates critical cell communication functions in normal development and may play a role in neoplasia. Here we describe a random mutagenesis approach, which we employed to study the molecular determinants of the EphA3/ephrin-A5 recognition. Selection and functional characterization of EphA3 point mutants with impaired ephrin-A5 binding from a yeast expression library defined three EphA3 surface areas that are essential for the EphA3/ephrin-A5 interaction. Two of these map to regions identified previously in the crystal structure of the homologous EphB2-ephrin-B2 complex as potential ligand/receptor interfaces. In addition, we identify a third EphA3/ephrin-A5 interface that falls outside the structurally characterized interaction domains. Functional analysis of EphA3 mutants reveals that all three Eph/ephrin contact areas are essential for the assembly of signaling-competent, oligomeric receptor-ligand complexes.