The SUN protein Mps3 is required for spindle pole body insertion into the nuclear membrane and nuclear envelope homeostasis

The budding yeast spindle pole body (SPB) is anchored in the nuclear envelope so that it can simultaneously nucleate both nuclear and cytoplasmic microtubules. During SPB duplication, the newly formed SPB is inserted into the nuclear membrane. The mechanism of SPB insertion is poorly understood but likely involves the action of integral membrane proteins to mediate changes in the nuclear envelope itself, such as fusion of the inner and outer nuclear membranes. Analysis of the functional domains of the budding yeast SUN protein and SPB component Mps3 revealed that most regions are not essential for growth or SPB duplication under wild-type conditions. However, a novel dominant allele in the P-loop region, MPS3-G186K, displays defects in multiple steps in SPB duplication, including SPB insertion, indicating a previously unknown role for Mps3 in this step of SPB assembly. Characterization of the MPS3-G186K mutant by electron microscopy revealed severe over-proliferation of the inner nuclear membrane, which could be rescued by altering the characteristics of the nuclear envelope using both chemical and genetic methods. Lipid profiling revealed that cells lacking MPS3 contain abnormal amounts of certain types of polar and neutral lipids, and deletion or mutation of MPS3 can suppress growth defects associated with inhibition of sterol biosynthesis, suggesting that Mps3 directly affects lipid homeostasis. Therefore, we propose that Mps3 facilitates insertion of SPBs in the nuclear membrane by modulating nuclear envelope composition.     

Recent increase in Nybelinia surmenicola prevalence and intensity in Pacific hake (Merluccius productus) off the United States west coast

A larval marine cestode was found in 82.0% of 834 Pacific hake (Merluccius productus) stomachs collected from 341 trawl stations along the United States west coast during the summers of 2008 and 2009. Morphology and DNA sequencing was used to identify the cestode as Nybelinia surmenicola. In an examination of 131 Pacific hake stomachs collected from the same region in 1999, N. surmenicola prevalence was 35.1%. The results from a general linear model suggested that their prevalence is influenced by year and latitude, Pacific hake size, and sex. Mean intensity of N. surmenicola in 2008-2009 was 20.22 (±1.13 SE) and was positively related to Pacific hake length and the latitude of collection. Year-1 Pacific hake (<27 cm length) had significantly lower prevalence and intensity of N. surmenicola compared to older and larger fish. Pacific hake collected south of Point Conception, California (32.5 to 35°N) had lower prevalence and intensity of N. surmenicola compared to those collected in northern latitudes (35.1 to 48.4°N). Higher N. surmenicola prevalence in Pacific hake in recent years suggests food-web fluctuations in the northern California current ecosystem caused by changes in ocean transport of zooplankton or pelagic fish distributions and warrants future monitoring as a metric for ecosystem change.     

Mapping the cofilin binding site on yeast G-actin by chemical cross-linking

Cofilin is a major cytoskeletal protein that binds to both monomeric actin (G-actin) and polymeric actin (F-actin) and is involved in microfilament dynamics. Although an atomic structure of the G-actin-cofilin complex does not exist, models of the complex have been built using molecular dynamics simulations, structural homology considerations, and synchrotron radiolytic footprinting data. The hydrophobic cleft between actin subdomains 1 and 3 and, alternatively, the cleft between actin subdomains 1 and 2 have been proposed as possible high-affinity cofilin binding sites. In this study, the proposed binding of cofilin to the subdomain 1/subdomain 3 region on G-actin has been probed using site-directed mutagenesis, fluorescence labeling, and chemical cross-linking, with yeast actin mutants containing single reactive cysteines in the actin hydrophobic cleft and with cofilin mutants carrying reactive cysteines in the regions predicted to bind to G-actin. Mass spectrometry analysis of the cross-linked complex revealed that cysteine 345 in subdomain 1 of mutant G-actin was cross-linked to native cysteine 62 on cofilin. A cofilin mutant that carried a cysteine substitution in the α3-helix (residue 95) formed a cross-link with residue 144 in actin subdomain 3. Distance constraints imposed by these cross-links provide experimental evidence for cofilin binding between actin subdomains 1 and 3 and fit a corresponding docking-based structure of the complex. The cross-linking of the N-terminal region of recombinant yeast cofilin to actin residues 346 and 374 with dithio-bis-maleimidoethane (12.4 Å) and via disulfide bond formation was also documented. This set of cross-linking data confirms the important role of the N-terminal segment of cofilin in interactions with G-actin.     

A new endogenous form of PYY isolated from canine ileum: Gly-extended PYY(1-36)

We purified and identified the peptide YY (PYY) forms present and determined their levels from a portion of the canine ileum directly adjacent to the cecum by a new extraction method designed to prevent and evaluate degradation of endogenous peptides. We used three reverse phase chromatography steps with radioimmunoassay of fractions for PYY-like-immunoreactivity (PYY-LI). The purified fractions underwent intact protein/peptide mass spectrometry identification and sequencing (i.e. "top-down" MS analysis). This analysis confirmed the identity of a new form of PYY, PYY(1-36)-Gly, which co-elutes with PYY(1-36)-NH(2) through all three of separation steps used. The PYY(1-36)-Gly form represents approximately 20% of the total PYY found in this region of the canine intestine. In addition, we also found that the PYY(3-36)-NH(2) form represents 6% of the total PYY in the canine ileo-cecal junction. The physiological implication of the Gly-extended form of PYY(1-36) warrants further investigation.     

Technologically indicative properties of straw fractions of flax, linseed (Linum usitatissimum L.) and fibre hemp (Cannabis sativa L.)

In this study of the behaviour of the fractions of unretted and frost-retted fibre straws in damp air, a production scale method to separate fibre and shive from fibre plants was introduced and tested on bast fibre plants (Linum usitatissimum L. and Cannabis sativa L.). The method consists of optional drying of stalks, unloading bales, milling the straws with a hammer mill, separating the fractions from air stream with a cyclone and finally separating fibres from shives with a screening drum. Fractions were characterized focusing on technologically indicative properties such as equilibrium moisture content, ash and microbiological quality. Unretted fractions of the bast fibre plant stem reached higher equilibrium moisture contents than the retted fractions, and hemp fibres absorbed more moisture from air than did the Linum fibres. In very humid air, all fractions began to lose weight due to moulding. The weight decrease during the first week was lower in frost-retted than in unretted fractions. The frost-retted fractions appeared to be more resistant to humidity in the short term. The total number of microbes and especially the numbers of yeasts and moulds can be used as a criterion of hygienic level. For green fractions, the mould level was similar in fibres and in shives, but frost-retted shives contained more moulds than the unretted shives. The mould content of a fraction had no direct correlation with the moulding tendency of the fraction. The ash contents of fibres were somewhat higher than those of shives, due to a probable soil contamination. Ash content did not have significant correlation with microbiological quality, although ash is a possible risk factor for hygienic quality. According to the results of this study it is highly important to study the quality of the production chain of bast fibre plants to ensure the quality of industrial products. From the producer's point of view, raw material with defined quality can be directed to the most suitable application. The behaviour of fractions in various ambient atmospheres, and other quality aspects such as hygienic level can be used as criteria for defining the most appropriate product applications.     

The relative contributions of seed and pollen dispersal to gene flow and genetic diversity in seedlings of a tropical palm

Seed and pollen dispersal shape patterns of gene flow and genetic diversity in plants. Pollen is generally thought to travel longer distances than seeds, but seeds determine the ultimate location of gametes. Resolving how interactions between these two dispersal processes shape microevolutionary processes is a long‐standing research priority. We unambiguously isolated the separate and combined contributions of these two dispersal processes in seedlings of the animal‐dispersed palm Oenocarpus bataua to address two questions. First, what is the spatial extent of pollen versus seed movement in a system characterized by long‐distance seed dispersal? Second, how does seed dispersal mediate seedling genetic diversity? Despite evidence of frequent long‐distance seed dispersal, we found that pollen moves much further than seeds. Nonetheless, seed dispersal ultimately mediates genetic diversity and fine‐scale spatial genetic structure. Compared to undispersed seedlings, seedlings dispersed by vertebrates were characterized by higher female gametic and diploid seedling diversity and weaker fine‐scale spatial genetic structure for female gametes, male gametes and diploid seedlings. Interestingly, the diversity of maternal seed sources at seed deposition sites (N _em) was associated with higher effective number of pollen sources (N _ep), higher effective number of parents (N _e) and weaker spatial genetic structure, whereas seed dispersal distance had little impact on these or other parameters we measured. These findings highlight the importance maternal seed source diversity (N _em) at frugivore seed deposition sites in driving emergent patterns of fine‐scale genetic diversity and structure.     

Identification of diamino chromone-2-carboxamides as MCHr1 antagonists with minimal hERG channel activity

A series of potent 2-carboxychromone-based melanin-concentrating hormone receptor 1 (MCHr1) antagonists were synthesized and evaluated for hERG (human Ether-a-go-go Related Gene) channel affinity and functional blockade. Basic dialkylamine-terminated analogs were found to weakly bind the hERG channel and provided marked improvement in a functional patch-clamp assay versus previously reported antagonists of the series.     

The effect of vapocoolant spray on pain due to intravenous cannulation in children: a randomized controlled trial

Background

Established noninvasive pharmacologic means of alleviating pain and anxiety in children undergoing intravenous cannulation are time-consuming, and thus impractical for routine use in the emergency department. Vapocoolant sprays provide transient skin anesthesia within seconds of application. We compared the effect of a new vapocoolant spray to placebo on pain due to intravenous cannulation in children.

Methods

In this double-blind randomized controlled trial, which we conducted between June 1 and Sept. 12, 2006, 80 children aged 6–12 years received either vapocoolant spray or placebo before cannulation. Children rated their pain using a 100-mm colour visual analogue scale. Secondary outcomes included success rate on first attempt at cannulation and pain ratings by the children''s parents, nurses and child life specialists.

Results

We found a modest but significant reduction in pain with the use of vapocoolant spray (mean difference 19 mm, 95% confidence interval [CI] 6–32 mm; p < 0.01). Cannulation on first attempt was more often successful with the use of vapocoolant spray (85.0%) than with placebo (62.5%) (mean difference 22.5%, 95% CI 3.2%–39.9%; p = 0.03). The number needed to treat to prevent 1 cannulation failure was 5 (95% CI 3–32). Parents (p = 0.04), nurses (p = 0.01) and child life specialists (p < 0.01) considered the children''s pain to be reduced with the use of vapocoolant spray.

Interpretation

The vapocoolant spray in our study quickly and effectively reduced pain due to intravenous cannulation in children and improved the success rate of cannulation. It is an important option to reduce childhood procedural pain in emergency situations, especially when time precludes traditional interventions.(http://ClinicalTrials.gov trial register no. {"type":"clinical-trial","attrs":{"text":"NCT00130650","term_id":"NCT00130650"}}NCT00130650.)Children frequently undergo intravenous cannulation in the emergency department. This painful procedure causes considerable stress and anxiety for children and their parents.¹ Failure to alleviate pain results in an uncooperative child, unsuccessful procedures (increased reattempts), prolonged procedure time and dissatisfaction with care for all involved.² Although pain may be reduced by behavioural and pharmacologic interventions (e.g., age-appropriate patient preparation, parental presence, distraction techniques, subcutaneous local anesthetics, topical anesthetic and systemic anesthetics), most of these preparations are impractical in nonelective settings because they are too time-consuming.^3–12 Novel delivery approaches and new medication compositions may decrease this time.^13,14 Vapocoolant sprays are rapid-acting alternatives to topical anesthetics. They provide transient anesthesia via evaporation-induced skin cooling, which reduces pain. Results from studies of earlier vapocoolant sprays indicated that they reduced pain due to vaccine injection in children and adults,^15–18 but not pain due to intravenous cannulation in children.^19,20 We sought to determine whether a new product, Pain Ease (Gebauer Company, Cleveland, Ohio), would reduce pain with intravenous cannulation in children.