181 research outputs found
Scanning mutagenesis of omega-atracotoxin-Hv1a reveals a spatially restricted epitope that confers selective activity against insect calcium channels
We constructed a complete panel of alanine mutants of the insect-specific calcium channel blocker omega-atracotoxin-Hv1a. Lethality assays using these mutant toxins identified three spatially contiguous residues, Pro(10), Asn(27), and Arg(35), that are critical for insecticidal activity against flies (Musca domestica) and crickets (Acheta domestica). Competitive binding assays using radiolabeled omega-atracotoxin-Hv1a and neuronal membranes prepared from the heads of American cockroaches (Periplaneta americana) confirmed the importance of these three residues for binding of the toxin to target calcium channels presumably expressed in the insect membranes. At concentrations up to 10 muM, omega-atracotoxin-Hv1a had no effect on heterologously expressed rat Ca(v)2.1, Ca(v)2.2, and Ca(v)1.2 calcium channels, consistent with the previously reported insect selectivity of the toxin. 30 muM omega-atracotoxin-Hv1a inhibited rat Ca-v currents by 10-34%, depending on the channel subtype, and this low level of inhibition was essentially unchanged when Asn(27) and Arg(35), which appears to be critical for interaction of the toxin with insect Ca-v channels, were both mutated to alanine. We propose that the spatially contiguous epitope formed by Pro(10), Asn(27), and Arg(35) confers specific binding to insect Ca-v channels and is largely responsible for the remarkable phyletic selectivity of omega-atracotoxin-Hv1a. This epitope provides a structural template for rational design of chemical insecticides that selectively target insect Ca-v channels
Control of Transcription by Cell Size
Cell size increases significantly with increasing ploidy. Differences in cell size and ploidy are associated with alterations in gene expression, although no direct connection has been made between cell size and transcription. Here we show that ploidy-associated changes in gene expression reflect transcriptional adjustment to a larger cell size, implicating cellular geometry as a key parameter in gene regulation. Using RNA-seq, we identified genes whose expression was altered in a tetraploid as compared with the isogenic haploid. A significant fraction of these genes encode cell surface proteins, suggesting an effect of the enlarged cell size on the differential regulation of these genes. To test this hypothesis, we examined expression of these genes in haploid mutants that also produce enlarged size. Surprisingly, many genes differentially regulated in the tetraploid are identically regulated in the enlarged haploids, and the magnitude of change in gene expression correlates with the degree of size enlargement. These results indicate a causal relationship between cell size and transcription, with a size-sensing mechanism that alters transcription in response to size. The genes responding to cell size are enriched for those regulated by two mitogen-activated protein kinase pathways, and components in those pathways were found to mediate size-dependent gene regulation. Transcriptional adjustment to enlarged cell size could underlie other cellular changes associated with polyploidy. The causal relationship between cell size and transcription suggests that cell size homeostasis serves a regulatory role in transcriptome maintenance.National Institutes of Health (U.S.) (grant GM035010)National Institutes of Health (U.S.) (grant GM040266
Functional Expression of Spider Neurotoxic Peptide Huwentoxin-I in E. coli
The coding sequence of huwentoxin-I, a neurotoxic peptide isolated from the venom of the Chinese spider Ornithoctonus huwena, was amplified by PCR using the cDNA library constructed from the spider venom glands. The cloned fragment was inserted into the expression vector pET-40b and transformed into the E. coli strain BL21 (DE3). The expression of a soluble fusion protein, disulfide interchange protein (DsbC)-huwentoxin-I, was auto-induced in the periplasm of E. coli in the absence of IPTG. After partial purification using a Ni-NTA column, the expressed fusion protein was digested using enterokinase to release heteroexpressed huwentoxin-I and was further purified using RP-HPLC. The resulting peptide was subjected to gel electrophoresis and mass spectrometry analysis. The molecular weight of the heteroexpressed huwentoxin-I was 3750.69, which is identical to that of the natural form of the peptide isolated from spider venom. The physiological properties of the heteroexpressed huwentoxin-I were further analyzed using a whole-cell patch clamp assay. The heteroexpressed huwentoxin-I was able to block currents generated by human Nav1.7 at an IC50 of 640 nmole/L, similar to that of the natural huwentoxin-I, which is 630 nmole/L
A Phylogeny and Timescale for the Evolution of Pseudocheiridae (Marsupialia: Diprotodontia) in Australia and New Guinea
Pseudocheiridae (Marsupialia: Diprotodontia) is a family of endemic Australasian arboreal folivores, more commonly known as ringtail possums. Seventeen extant species are grouped into six genera (Pseudocheirus, Pseudochirulus, Hemibelideus, Petauroides, Pseudochirops, Petropseudes). Pseudochirops and Pseudochirulus are the only genera with representatives on New Guinea and surrounding western islands. Here, we examine phylogenetic relationships among 13 of the 17 extant pseudocheirid species based on protein-coding portions of the ApoB, BRCA1, ENAM, IRBP, Rag1, and vWF genes. Maximum parsimony, maximum likelihood, and Bayesian methods were used to estimate phylogenetic relationships. Two different relaxed molecular clock methods were used to estimate divergence times. Bayesian and maximum parsimony methods were used to reconstruct ancestral character states for geographic provenance and maximum elevation occupied. We find robust support for the monophyly of Pseudocheirinae (Pseudochirulus + Pseudocheirus), Hemibelidinae (Hemibelideus + Petauroides), and Pseudochiropsinae (Pseudochirops + Petropseudes), respectively, and for an association of Pseudocheirinae and Hemibelidinae to the exclusion of Pseudochiropsinae. Within Pseudochiropsinae, Petropseudes grouped more closely with the New Guinean Pseudochirops spp. than with the Australian Pseudochirops archeri, rendering Pseudochirops paraphyletic. New Guinean species belonging to Pseudochirops are monophyletic, as are New Guinean species belonging to Pseudochirulus. Molecular dates and ancestral reconstructions of geographic provenance combine to suggest that the ancestors of extant New Guinean Pseudochirops spp. and Pseudochirulus spp. dispersed from Australia to New Guinea ∼12.1–6.5 Ma (Pseudochirops) and ∼6.0–2.4 Ma (Pseudochirulus). Ancestral state reconstructions support the hypothesis that occupation of high elevations (>3000 m) is a derived feature that evolved on the terminal branch leading to Pseudochirops cupreus, and either evolved in the ancestor of Pseudochirulus forbesi, Pseudochirulus mayeri, and Pseudochirulus caroli, with subsequent loss in P. caroli, or evolved independently in P. mayeri and P. forbesi. Divergence times within the New Guinean Pseudochirops clade are generally coincident with the uplift of the central cordillera and other highlands. Diversification within New Guinean Pseudochirulus occurred in the Plio-Pleistocene after the establishment of the Central Range and other highlands
Randomised clinical study: inulin short-chain fatty acid esters for targeted delivery of short-chain fatty acids to the human colon
SUMMARY Background Short-chain fatty acids (SCFA) produced through fermentation of nondigestible carbohydrates by the gut microbiota are associated with positive metabolic effects. However, well-controlled trials are limited in humans
Effect of age, sex and gender on pain sensitivity: A narrative review
© 2017 Eltumi And Tashani. Introduction: An increasing body of literature on sex and gender differences in pain sensitivity has been accumulated in recent years. There is also evidence from epidemiological research that painful conditions are more prevalent in older people. The aim of this narrative review is to critically appraise the relevant literature investigating the presence of age and sex differences in clinical and experimental pain conditions. Methods: A scoping search of the literature identifying relevant peer reviewed articles was conducted on May 2016. Information and evidence from the key articles were narratively described and data was quantitatively synthesised to identify gaps of knowledge in the research literature concerning age and sex differences in pain responses. Results: This critical appraisal of the literature suggests that the results of the experimental and clinical studies regarding age and sex differences in pain contain some contradictions as far as age differences in pain are concerned. While data from the clinical studies are more consistent and seem to point towards the fact that chronic pain prevalence increases in the elderly findings from the experimental studies on the other hand were inconsistent, with pain threshold increasing with age in some studies and decreasing with age in others. Conclusion: There is a need for further research using the latest advanced quantitative sensory testing protocols to measure the function of small nerve fibres that are involved in nociception and pain sensitivity across the human life span. Implications: Findings from these studies should feed into and inform evidence emerging from other types of studies (e.g. brain imaging technique and psychometrics) suggesting that pain in the older humans may have unique characteristics that affect how old patients respond to intervention
Using direct observations on multiple occasions to measure household food availability among low-income Mexicano residents in Texas colonias
BACKGROUND:
It has been recognized that the availability of foods in the home are important to nutritional health, and may influence the dietary behavior of children, adolescents, and adults. It is therefore important to understand food choices in the context of the household setting. Considering their importance, the measurement of household food resources becomes critical.Because most studies use a single point of data collection to determine the types of foods that are present in the home, which can miss the change in availability within a month and when resources are not available, the primary objective of this pilot study was to examine the feasibility and value of conducting weekly in-home assessments of household food resources over the course of one month among low-income Mexicano families in Texas colonias.
METHODS:
We conducted five in-home household food inventories over a thirty-day period in a small convenience sample; determined the frequency that food items were present in the participating households; and compared a one-time measurement with multiple measurements.After the development and pre-testing of the 252-item culturally and linguistically- appropriate household food inventory instrument that used direct observation to determine the presence and amount of food and beverage items in the home (refrigerator, freezer, pantry, elsewhere), two trained promotoras recruited a convenience sample of 6 households; administered a baseline questionnaire (personal info, shopping habits, and food security); conducted 5 in-home assessments (7-day interval) over a 30-day period; and documented grocery shopping and other food-related activities within the previous week of each in-home assessment. All data were collected in Spanish. Descriptive statistics were calculated for mean and frequency of sample characteristics, food-related activities, food security, and the presence of individual food items. Due to the small sample size of the pilot data, the Friedman Test and Kendall's W were used to assess the consistency of household food supplies across multiple observations.
RESULTS:
Complete data were collected from all 6 Mexicano women (33.2y +/- 3.3; 6.5 +/- 1.5 adults/children in household (HH); 5 HH received weekly income; and all were food insecure. All households purchased groceries within a week of at least four of the five assessments. The weekly presence and amounts of fresh and processed fruits and vegetables, dairy, meats, breads, cereals, beverages, and oils and fats varied. Further, the results revealed the inadequacy of a one-time measurement of household food resources, compared with multiple measures. The first household food inventory as a one-time measure would have mistakenly identified at least one-half of the participant households without fresh fruit, canned vegetables, dairy, protein foods, grains, chips, and sugar-sweetened beverages.
CONCLUSIONS:
This study highlights the value of documenting weekly household food supplies, especially in households where income resources may be more volatile. Clearly, the data show that a single HFI may miss the changes in availability--presence and amount--that occur among low-income Mexicano households who face challenges that require frequent purchase of foods and beverages. Use of multiple household food inventories can inform the development and implementation of nutrition-related policies and culturally sensitive nutrition education programs
Estudo comparativo entre os sincrânios de Otaria byronia e Arctocephalus australis (Pinnipedia, Otariidae)
Is there an interaction between perceived direction and perceived aspect ratio in stereoscopic vision?
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