Structural basis of PxxDY motif recognition in SH3 binding

We have determined the solution structure of epidermal growth factor receptor pathway substrate 8 (Eps8) L1 Src homology 3 (SH3) domain in complex with the PPVPNPDYEPIR peptide from the CD3ε cytoplasmic tail. Our structure reveals the distinct structural features that account for the unusual specificity of the Eps8 family SH3 domains for ligands containing a PxxDY motif instead of canonical PxxP ligands. The CD3ε peptide binds Eps8L1 SH3 in a class II orientation, but neither adopts a polyproline II helical conformation nor engages the first proline-binding pocket of the SH3 ligand binding interface. Ile531 of Eps8L1 SH3, instead of Tyr or Phe residues typically found in this position in SH3 domains, renders this hydrophobic pocket smaller and nonoptimal for binding to conventional PxxP peptides. A positively charged arginine at position 512 in the n-Src loop of Eps8L1 SH3 plays a key role in PxxDY motif recognition by forming a salt bridge to D7 of the CD3ε peptide. In addition, our structural model suggests a hydrogen bond between the hydroxyl group of the aromatic ring of Y8 and the carboxyl group of E496, thus explaining the critical role of the PxxDY motif tyrosine residue in binding to Eps8 family SH3. These finding have direct implications also for understanding the atypical binding specificity of the amino-terminal SH3 of the Nck family proteins.     

Growth of reindeer lichens and effects of reindeer grazing on ground cover vegetation in a Scots pine forest and a subarctic heathland in Finnish Lapland

Reindeer lichens are an important component of northern ecosystems. The aim of this study was to measure the growth rate of terricolous lichens as it is a key parameter involved in productivity of these ecosystems and an important part of lichen tolerance to reindeer grazing. Furthermore, the natural succession and the long-term effects of reindeer grazing on lichen community characteristics in two contrasting habitats were investigated as well as the interactions between lichen cover and mosses and vascular plants. Biomass and coverage measurements were conducted in a lichen woodland and in a subarctic heath with grazed and ungrazed areas in northern Finland. Measurements spanning over 13 yr of undisturbed development show that the growth rate of Cladina stellaris can be as high as >0.17 g g⁻¹ produced annually, although in average growth rates were much lower. During the succession of ground vegetation, C. stellaris , C. rangiferina , C. mitis and Cetraria nivalis increased in biomass in fenced areas and were reduced most in biomass by reindeer in unprotected areas. Reindeer grazing and trampling seem to change the vegetation towards a type that is dominated by small dwarf shrubs, bare soil and minute-cup lichens ( Cladonia spp.). Removing the lichen layer by reindeer may reduce natural regeneration of pine trees as implied by increasing numbers of pine seedlings with increasing lichen cover.     

An intraresidual i(HCA)CO(CA)NH experiment for the assignment of main-chain resonances in <Superscript>15</Superscript>N, <Superscript>13</Superscript>C labeled proteins

An improved pulse sequence, intraresidual i(HCA)CO(CA)NH, is described for establishing solely ¹³C′(i), ¹⁵N(i), ¹H^N(i) connectivities in uniformly ¹⁵N/¹³C-labeled proteins. In comparison to the “out-and-back” style intra-HN(CA)CO experiment, the new pulse sequence offers at least two-fold higher experimental resolution in the ¹³C′ dimension and on average 1.6 times higher sensitivity especially for residues in α-helices. Performance of the new experiment was tested on a small globular protein ubiquitin and an intrinsically unfolded 110-residue cancer/testis antigen CT16/PAGE5. Use of intraresidual i(HCA)CO(CA)NH experiment in combination with the established HNCO experiment was crucial for the assignment of highly disordered CT16.     

BODIPY-cholesterol: a new tool to visualize sterol trafficking in living cells and organisms

Analysis of sterol distribution and transport in living cells has been hampered by the lack of bright, photostable fluorescent sterol derivatives that closely resemble cholesterol. In this study, we employed atomistic simulations and experiments to characterize a cholesterol compound with fluorescent boron dipyrromethene difluoride linked to sterol carbon-24 (BODIPY-cholesterol). This probe packed in the membrane and behaved similarly to cholesterol both in normal and in cholesterol-storage disease cells and with trace amounts allowed the visualization of sterol movement in living systems. Upon injection into the yolk sac, BODIPY-cholesterol did not disturb zebrafish development and was targeted to sterol-enriched brain regions in live fish. We conclude that this new probe closely mimics the membrane partitioning and trafficking of cholesterol and, because of its excellent fluorescent properties, enables the direct monitoring of sterol movement by time-lapse imaging using trace amounts of the probe. This is, to our knowledge, the first cholesterol probe that fulfills these prerequisites.     

Molecular phylogeny of the speciose vole genus Microtus (Arvicolinae, Rodentia) inferred from mitochondrial DNA sequences

Voles of the genus Microtus represent one of the most speciose mammalian genera in the Holarctic. We established a molecular phylogeny for Microtus to resolve contentious issues of systematic relationships and evolutionary history in this genus. A total of 81 specimens representing ten Microtus species endemic to Europe as well as eight Eurasian, six Asian and one Holarctic species were sequenced for the entire cytochrome b gene (1140 bp). A further 25 sequences were retrieved from GenBank, providing data on an additional 23, mainly Nearctic, Microtus species. Phylogenetic analysis of these 48 species generated four well-supported monophyletic lineages. The genus Chionomys, snow voles, formed a distinct and well-supported lineage separate from the genus Microtus. The subgenus Microtus formed the strongest supported lineage with two sublineages displaying a close relationship between the arvalis species group (common voles) and the socialis species group (social voles). Monophyly of the Palearctic pitymyid voles, subgenus Terricola, was supported, and this subgenus was also subdivided into two monophyletic species groups. Together, these groupings clarify long-standing taxonomic uncertainties in Microtus. In addition, the "Asian" and the Nearctic lineages reported previously were identified although the latter group was not supported. However, relationships among the main Microtus branches were not resolved, suggesting a rapid and potentially simultaneous radiation of a widespread ancestor early in the history of the genus. This and subsequent radiations discernible in the cytochrome b phylogeny, show the considerable potential of Microtus for analysis of historical and ecological determinants of speciation in small mammals. It is evident that speciation is an ongoing process in the genus and that the molecular data provides a vital insight into current species limits as well as cladogenic events of the past.     

Multigenic and morphometric differentiation of ground squirrels (Spermophilus, Scuiridae, Rodentia) in Turkey, with a description of a new species

This study reports on the molecular phylogenetics of ground squirrels, genus Spermophilus, in Turkey using cytochrome b (1140bp), part of the D-loop and flanking tRNAs (572bp), X chromosome (867-1051bp) and Y chromosome (983-989bp) DNA sequences. Individuals also were characterized by karyotype and with geometric morphometric analyses of mandibles and skulls. Two hundred fourteen individuals from 91 localities were studied. All the data support the recognition of a new species in SW Anatolia: the Taurus ground squirrel Spermophilus taurensis sp. nov. The new species has a small distribution in the Taurus Mountains in an area that is a hotspot for biodiversity. Molecular clock analysis suggests that the new species diverged from the European ground squirrel, Spermophilus citellus, about 2.5 million years ago and that the ancestor of these two species diverged from the widespread Anatolian ground squirrel, Spermophilus xanthoprymnus, about 5 million years ago. Morphometric differentiation in skull and mandible shape among the three species is incomplete, but statistically significant. S. xanthoprymnus is subdivided into five cytochrome b phylogroups and we use these data to infer the location of glacial refugia where the species lived during the last glacial maximum. This study illustrates the potential of combined molecular and morphometric studies to uncover new Anatolian species and to reconstruct their phylogeographic history. The new species is important for squirrel taxonomy and for understanding Eurasian mammal evolution.     

Perennial polypores as indicators of annual and red-listed polypores

Many polypores are specialized in their requirements for substrate and environment, and they have been suggested to indicate the continuity of coarse woody debris or naturalness of a forest stand. However, the use of polypores as indicators of conservation value is restricted by the temporally limited appearance of annual fruit bodies. We studied whether the species richness of perennial polypores (perennials) can be used to predict the species richness of annual or annual red-listed polypores (annuals). Our data included 1471 separate datasets (sample plots or larger inventoried areas) in different parts of Finland and Russian Karelia, ranging from the southern to northern boreal zone. At the large scale (the whole area) the number of perennials explained about 70% of the variation in the number of annuals, and about 67% in the number of red-listed annuals. A minimum set of 40–60 perennial occurrences gave a reliable estimate on the species richness of annuals, and 60–80 occurrences on the species richness of red-listed annuals. The richness of perennials predicted the richness of annuals and, in particular, richness of red-listed annuals, better than the size of inventoried area. According to our results, perennial polypores can be used as a surrogate for overall polypore species richness in natural and seminatural boreal forests, but the predictive power is weaker in managed forests. In addition, the relationship between the perennial and annual species seems to differ in different vegetation zones, management types and forest types. Due to this variation direct application of the indicator values derived from different vegetation zones and management or forest types are not recommended. Since perennials are easier to identify than annuals, detectable throughout the year, and have much smaller year-to-year variation, their use as an indicator group seems to offer advantages regarding the timing and cost-efficiency of inventories.     

The Impact of Hypnotic Suggestions on Reaction Times in Continuous Performance Test in Adults with ADHD and Healthy Controls

Attention is one of the key factors in both hypnotic processes and patients with ADHD. In addition, the brain areas associated with hypnosis and ADHD overlap in many respects. However, the use of hypnosis in ADHD patients has still received only minor attention in research. The main purpose of the present work was to investigate whether hypnosis and hypnotic suggestions influence the performance of adult ADHD (n = 27) and control participants (n = 31) in the continuous performance test (CPT). The hypnotic susceptibility of the participants was measured by the Harvard Group Scale of Hypnotic Susceptibility (HGSHS:A) and the attentional task was a three minute long auditory version of the CPT. The CPT task was administered four times: before hypnosis (CPT1), after a hypnotic induction (CPT2), after suggestions about speed and accuracy (CPT3), and after the termination of hypnosis (CPT4). The susceptibility of the groups measured by HGSHS:A did not differ. There was a statistically significant decrease in reaction times in both ADHD and control groups between CPT2 and CPT3. The differences between CPT1 and CPT2, even though non-significant, were different in the two groups: in the ADHD group reaction times decreased whereas in the control group they increased. Both groups made very few errors in the short CPT. This study indicates that hypnotic suggestions have an effect on reaction times in the sustained attention task both in adult ADHD patients and control subjects. The theoretical and clinical implications are discussed.