A computational and experimental approach toward a priori identification of alternatively spliced exons

Alternative splicing is a powerful means of regulating gene expression and enhancing protein diversity. In fact, the majority of metazoan genes encode pre-mRNAs that are alternatively spliced to produce anywhere from two to tens of thousands of mRNA isoforms. Thus, an important part of determining the complete proteome of an organism is developing a catalog of all mRNA isoforms. Alternatively spliced exons are typically identified by aligning EST clusters to reference mRNAs or genomic DNA. However, this approach is not useful for genomes that lack robust EST coverage, and tools that enable accurate prediction of alternatively spliced exons would be extraordinarily useful. Here, we use comparative genomics to identify, and experimentally verify, potential alternative exons based solely on their high degree of conservation between Drosophila melanogaster and D. pseudoobscura. At least 40% of the exons that fit our prediction criteria are in fact alternatively spliced. Thus, comparative genomics can be used to accurately predict certain classes of alternative exons without relying on EST data.     

Comparison of changes in toxicity during growth of Anabaena circinalis (cyanobacteria) determined by mouse neuroblastoma bioassay and HPLC

Changes in toxicity were studied during batch culture of a strain ofAnabaena circinalis (AWQC ANA-311F) producing paralyticshellfish toxins (PSTs). PSTs were extracted from cells and culture mediumduring the culture period of 29 days. Samples were analysed by HPLC withpost-column oxidation coupled to fluorescence detection, and tested with themouse neuroblastoma bioassay (MNB). Seven PSTs were detected, but only fivewerequantified. Based on chemical analysis of PSTs by HPLC, atheoretical saxitoxin concentration related to mouse toxicity wascalculated. Neuroblastoma cell survival was measured during the MNB andcorrelated to total toxicity of the A. circinalis samples.Comparison was made between the changes in total toxicity shown by MNB and thetheoretical saxitoxin concentration shown by HPLC. There was asignificant positive correlation between the two in the culture medium, but notthe cells.     

The evolution of competitive strategies in annual plants

Annual plants are common in disturbed habitats. It is frequently assumed that because these habitats often have low-plant density, competition is not important in shaping the ecological strategies of annual plants. We test for competitive strategies in genotypes of the short-lived annual plant Arabidopsis thaliana. Genotypes were grown in treatments with or without conspecific competitors. We measured size at reproduction and fitness (fruit production) at final development. We estimated competitive ability in each genotype at first reproduction (the ability to maintain size in the presence of competitors) and at final development (the ability to maintain fruit production in the presence of competitors). Genotypes showed relatively high competitive ability measured as fruit production at final development, but most genotypes had low competitive ability measured as size at reproduction. Our results suggest that competition has been important in the evolution of strategies in these genotypes but vegetative size is not a strong predictor of competitive ability. Rather, competitive ability is determined by the capacity to reproduce efficiently in the presence of competitors. The competitive strategies expressed across these genotypes suggest that competition has been a selective force in these plants, and that a fast life history is not equivalent to an “r-strategy.”     

Rheumatic Heart Disease-Attributable Mortality at Ages 5–69 Years in Fiji: A Five-Year,National, Population-Based Record-Linkage Cohort Study

BackgroundRheumatic heart disease (RHD) is considered a major public health problem in developing countries, although scarce data are available to substantiate this. Here we quantify mortality from RHD in Fiji during 2008–2012 in people aged 5–69 years.ConclusionsRheumatic heart disease is a leading cause of premature death as well as an important economic burden in this setting. Age-standardised death rates are more than twice those reported in current global estimates. Linkage of routine data provides an efficient tool to better define the epidemiology of neglected diseases.     

Conditions Associated with Circulating Tumor-Associated Folate Receptor 1 Protein in Healthy Men and Women

Background

Serum concentrations of the tumor-associated folate receptor 1 (FOLR1) protein may be a marker for early cancer detection, yet concentrations have also been detected in cancer-free women. We investigated the conditions associated with circulating FOLR1 protein in healthy individuals and sought to clarify the range of normal serum values.

Methods

Sera of cancer-free men and women (N = 60) enrolled in a population-based cohort study in Alberta, Canada were analyzed for FOLR1 protein using an electrochemical luminescence immunoassay. Dietary, lifestyle, medical and reproductive history information was collected by questionnaires. Differences in serum FOLR1 concentrations between groups were assessed by non-parametric tests, and predictors of serum FOLR1 concentrations were estimated using multivariable linear regression.

Results

Median serum FOLR1 concentration was higher in women (491 pg/ml, range = 327–693 pg/ml) than in men (404 pg/ml, range = 340–682 pg/ml), P = 0.001. FOLR1 concentration was also positively associated with vitamin A intake (P = 0.02), and showed positive trends with age and with oral contraceptive hormone use among women and an inverse trend with body mass index. All variables examined explained almost half of the variation in serum FOLR1 (model R² = 0.44, P = 0.04); however, the retention of gender (P = 0.003) and vitamin A intake (P = 0.03) together explained 20% (P = 0.001) of serum FOLR1 variation. No other predictor was significant at P<0.05.

Conclusions

The positive association between serum FOLR1 concentration and female gender independent of an age effect suggests caution against statements to exploit serum FOLR1 for early cancer detection without further understanding the biological underpinnings of these observations. Serum FOLR1 concentrations may be influenced by the steroid retinoic acid (vitamin A) but do not appear to be associated with folate nutritional status. These findings require confirmation in larger independent studies.     

Loss of DNA mismatch repair imparts a selective advantage in planarian adult stem cells

Lynch syndrome (LS) leads to an increased risk of early-onset colorectal and other types of cancer and is caused by germline mutations in DNA mismatch repair (MMR) genes. Loss of MMR function results in a mutator phenotype that likely underlies its role in tumorigenesis. However, loss of MMR also results in the elimination of a DNA damage-induced checkpoint/apoptosis activation barrier that may allow damaged cells to grow unchecked. A fundamental question is whether loss of MMR provides pre-cancerous stem cells an immediate selective advantage in addition to establishing a mutator phenotype. To test this hypothesis in an in vivo system, we utilized the planarian Schmidtea mediterranea which contains a significant population of identifiable adult stem cells. We identified a planarian homolog of human MSH2, a MMR gene which is mutated in 38% of LS cases. The planarian Smed-msh2 is expressed in stem cells and some progeny. We depleted Smed-msh2 mRNA levels by RNA-interference and found a striking survival advantage in these animals treated with a cytotoxic DNA alkylating agent compared to control animals. We demonstrated that this tolerance to DNA damage is due to the survival of mitotically active, MMR-deficient stem cells. Our results suggest that loss of MMR provides an in vivo survival advantage to the stem cell population in the presence of DNA damage that may have implications for tumorigenesis.     

RNA structure and the mechanisms of alternative splicing

Alternative splicing is a widespread means of increasing protein diversity and regulating gene expression in eukaryotes. Much progress has been made in understanding the proteins involved in regulating alternative splicing, the sequences they bind to, and how these interactions lead to changes in splicing patterns. However, several recent studies have identified other players involved in regulating alternative splicing. A major theme emerging from these studies is that RNA secondary structures play an under appreciated role in the regulation of alternative splicing. This review provides an overview of the basic aspects of splicing regulation and highlights recent progress in understanding the role of RNA secondary structure in this process.     

Engineered water level management facilitates recruitment of non-native common carp, Cyprinus carpio, in a regulated lowland river

River regulation can advantage non-native aquatic biota at the expense of native species. Nevertheless, flow regulating structures are sometimes used with the aim of achieving positive environmental outcomes in aquatic ecosystems. In the lower River Murray, Australia, drought-induced water level recession and acid sulfate soil exposure prompted the construction of an earthen levee, isolating a section of river channel (the Goolwa weir pool (GWP)) within which water levels were managed to mitigate a risk of water body acidification. The present study aimed to determine the impact of water level management on the fish community by investigating variation in species abundance and recruitment between sites subject to water level management in the GWP and unmanaged sites in Lake Alexandrina. Prior to levee construction, in August 2009, the abundance of the non-native common carp was similar in the GWP and Lake Alexandrina. Following water level management, in December 2009 and April 2010, the abundance of common carp in the GWP was approximately 1000 and 250 times greater than abundance in Lake Alexandrina, as a result of recruitment of young-of-year fish. No native freshwater species were significantly more abundant in the GWP in August 2009, December 2009 or April 2010. The results of this study suggest that the isolation of a river reach and a managed rise in water level facilitated spawning and recruitment of a non-native fish species. As such, the ecological benefits and risks of restoration and mitigation projects that involve the construction of flow regulating structures and water level management should be carefully considered.     

Bactericidal Efficacy of Oxidized Silver against Biofilms Formed by Curtobacterium flaccumfaciens pv. flaccumfaciens

Bacterial wilt is a re-emerging disease on dry bean and can affect many other crop species within the Fabaceae. The causal agent, Curtobacterium flaccumfaciens pv. flaccumfaciens (CFF), is a small, Gram-positive, rod-shaped bacterium that is seed-transmitted. Infections in the host become systemic, leading to wilting and economic loss. Clean seed programs and bactericidal seed treatments are two critical management tools. This study characterizes the efficacies of five bactericidal chemicals against CFF. It was hypothesized that this bacterium was capable of forming biofilms, and that the cells within biofilms would be more tolerant to bactericidal treatments. The minimum biocide eradication concentration assay protocol was used to grow CFF biofilms, expose the biofilms to bactericides, and enumerate survivors compared to a non-treated control (water). Streptomycin and oxysilver bisulfate had EC₉₅ values at the lowest concentrations and are likely the best candidates for seed treatment products for controlling seed-borne bacterial wilt of bean. The results showed that CFF formed biofilms during at least two phases of the bacterial wilt disease cycle, and the biofilms were much more difficult to eradicate than their planktonic counterparts. Overall, biofilm formation by CFF is an important part of the bacterial wilt disease cycle in dry edible bean and antibiofilm bactericides such as streptomycin and oxysilver bisulfate may be best suited for use in disease management.     

Aggressive interactions between three species of freshwater crayfish of the genus Cherax (Decapoda: Parastacidae)

The freshwater crayfish, Cherax destructor Clark, native to southeastern Australia, was first introduced to farm dams in southwestern Western Australia in 1932. The geographic range of the crayfish in Western Australia has increased substantially since then, and in recent years it has become established in natural waterways where it co-occurs with species of freshwater crayfish endemic to southwestern Australia, Cherax cainii and Cherax quinquecarinatus. The potential for competitive exclusion of these endemic species by C. destructor was investigated through laboratory experiments measuring aggressive behaviour. Body mass and species were found to be important factors governing aggressive dominance between C. cainii and C. destructor, with C. cainii winning significantly more interactions only when they were larger in body mass than their opponent. In trials between C. quinquecarinatus and C. destructor of similar body mass, there was no difference between the number of interactions 'won' by the two species. The implications for natural populations are discussed.