Seemingly neutral polymorphic variants may confer immunity to splicing-inactivating mutations: a synonymous SNP in exon 5 of MCAD protects from deleterious mutations in a flanking exonic splicing enhancer

The idea that point mutations in exons may affect splicing is intriguing and adds an additional layer of complexity when evaluating their possible effects. Even in the best-studied examples, the molecular mechanisms are not fully understood. Here, we use patient cells, model minigenes, and in vitro assays to show that a missense mutation in exon 5 of the medium-chain acyl-CoA dehydrogenase (MCAD) gene primarily causes exon skipping by inactivating a crucial exonic splicing enhancer (ESE), thus leading to loss of a functional protein and to MCAD deficiency. This ESE functions by antagonizing a juxtaposed exonic splicing silencer (ESS) and is necessary to define a suboptimal 3′ splice site. Remarkably, a synonymous polymorphic variation in MCAD exon 5 inactivates the ESS, and, although this has no effect on splicing by itself, it makes splicing immune to deleterious mutations in the ESE. Furthermore, the region of MCAD exon 5 that harbors these elements is nearly identical to the exon 7 region of the survival of motor neuron (SMN) genes that contains the deleterious silent mutation in SMN2, indicating a very similar and finely tuned interplay between regulatory elements in these two genes. Our findings illustrate a mechanism for dramatic context-dependent effects of single-nucleotide polymorphisms on gene-expression regulation and show that it is essential that potential deleterious effects of mutations on splicing be evaluated in the context of the relevant haplotype.     

Raver1 is an integral component of muscle contractile elements

Raver1, a ubiquitously expressed protein, was originally identified as a ligand for metavinculin, the muscle-specific isoform of the microfilament-associated protein vinculin. The protein resides primarily in the nucleus, where it colocalises and may interact with polypyrimidine-tract-binding protein, which is involved in alternative splicing processes. During skeletal muscle differentiation, raver1 translocates to the cytoplasm and eventually targets the Z-line of sarcomeres. Here, it colocalises with metavinculin, vinculin and alpha-actinin, all of which have biochemically been identified as raver1 ligands. To obtain more information about the potential role of raver1 in muscle structure and function, we have investigated its distribution and fine localisation in mouse striated and smooth muscle, by using three monoclonal antibodies that recognise epitopes in different regions of the raver1 protein. Our immunofluorescence and immunoelectron-microscopic results indicate that the cytoplasmic accumulation of raver1 is not confined to skeletal muscle but also occurs in heart and smooth muscle. Unlike vinculin and metavinculin, cytoplasmic raver1 is not restricted to costameres but additionally represents an integral part of the sarcomere. In isolated myofibrils and in ultrathin sections of skeletal muscle, raver1 has been found concentrated at the I-Z-I band. A minor fraction of raver1 is present in the nuclei of all three types of muscle. These data indicate that, during muscle differentiation, raver1 might link gene expression with structural functions of the contractile machinery of muscle. This work was supported by grants from the Swiss National Science Foundation and the M.E. Müller Foundation (to C.A.S.) and the Deutsche Forschungsgemeinschaft (to S.I. and B.M.J.) and from the Fonds der Chemischen Industrie (to B.M.J.). A.Z. was the recipient of a G. Lichtenberg fellowship, within an International Graduate College funded by the State of Lower Saxony, Germany.     

The influence of enzymatic treatment on wool fibre properties using PEG-modified proteases

The main contribution of the presented work was to introduce the use of proteases modified with the soluble polymer polyethylene glycol (PEG) in the bio-finishing process of wool fibres, to target enzyme action to the outer parts of wool fibres, i.e. to avoid the diffusion and consequent destroying of the inner parts of the wool fibre structure, in the case of native proteases using.

Different proteolytic enzymes from Bacillus lentus and Bacillus subtilis in native and PEG-modified forms were investigated and their influence on the modification of wool fibres morphology surface, chemical structure, as well as the hydrolysis of wool proteins, the physico-mechanical properties, and the sorption properties of 1:2 metal complex dye during dyeing were studied. SEM images of wool fibres confirmed smoother and cleaner fibre surfaces without fibre damages using PEG-modified proteases. Modified enzyme products have a benefit effect on the wool fibres felting behaviours (14%) in the case when PEG-modified B. lentus is used, without markedly fibre damage expressed by tensile strength and weight loss of the fibre. Meanwhile the dye exhaustion showed slower but comparable level of dye uptake at the end of the dyeing.     

The -11A of promoter DNA and two conserved amino acids in the melting region of sigma70 both directly affect the rate limiting step in formation of the stable RNA polymerase-promoter complex, but they do not necessarily interact

Formation of the stable, strand separated, ‘open’ complex between RNA polymerase and a promoter involves DNA melting of approximately 14 base pairs. The likely nucleation site is the highly conserved −11A base in the non-template strand of the −10 promoter region. Amino acid residues Y430 and W433 on the σ⁷⁰ subunit of the RNA polymerase participate in the strand separation. The roles of −11A and of the Y430 and W433 were addressed by employing synthetic consensus promoters containing base analog and other substitutions at −11 in the non-template strand, and σ⁷⁰ variants bearing amino acid substitutions at positions 430 and 433. Substitutions for −11A and for Y430 and W433 in σ⁷⁰ have small or no effects on formation of the initial RNA polymerase-promoter complex, but exert their effects on subsequent steps on the way to formation of the open complex. As substitutions for Y430 and W433 also affect open complex formation on promoter DNA lacking the −11A base, it is concluded that these amino acid residues have other (or additional) roles, not involving the −11A. The effects of the substitutions at −11A of the promoter and Y430 and W433 of σ⁷⁰ are cumulative.     

Male age and its association with reproductive traits in captive and wild house sparrows

Evolutionary theory predicts that females seek extra‐pair fertilizations from high‐quality males. In socially monogamous bird species, it is often old males that are most successful in extra‐pair fertilizations. Adaptive models of female extra‐pair mate choice suggest that old males may produce offspring of higher genetic quality than young males because they have proven their survivability. However, old males are also more likely to show signs of reproductive senescence, such as reduced sperm quality. To better understand why old males account for a disproportionally large number of extra‐pair offspring and what the consequences of mating with old males are, we compared several sperm traits of both captive and wild house sparrows, Passer domesticus. Sperm morphological traits and cloacal protuberance volume (a proxy for sperm load) of old and young males did not differ substantially. However, old males delivered almost three times more sperm to the female's egg than young males. We discuss the possibility of a post‐copulatory advantage for old over young males and the consequences for females mated with old males.     

The Relevance of Circular Economy Practices to the Sustainable Development Goals

This paper identifies the extent to which circular economy (CE) practices are relevant for the implementation of the Sustainable Development Goals (SDGs). The results of a literature review and a matching exercise to determine the relationship between CE practices and SDG targets show that CE practices, potentially, can contribute directly to achieving a significant number of SDG targets. The strongest relationships exist between CE practices and the targets of SDG 6 (Clean Water and Sanitation), SDG 7 (Affordable and Clean Energy), SDG 8 (Decent Work and Economic Growth), SDG 12 (Responsible Consumption and Production), and SDG 15 (Life on Land). The paper also explores synergies that can be created through CE practices among several of the SDG targets. Furthermore, it identifies several potential trade‐offs between targets for decent work, safe working environments, human health and current CE practices relating to recycling of municipal waste, e‐waste and wastewater, and provides suggestions how these can be overcome. The paper concludes that CE practices can be applied as a “toolbox” and specific implementation approaches for achieving a sizeable number of SDG targets. Further empirical research is necessary to determine which specific types of partnerships and means of implementation are required to apply CE practices in the SDG context.     

Violation of an Evolutionarily Conserved Immunoglobulin Diversity Gene Sequence Preference Promotes Production of dsDNA-Specific IgG Antibodies

Variability in the developing antibody repertoire is focused on the third complementarity determining region of the H chain (CDR-H3), which lies at the center of the antigen binding site where it often plays a decisive role in antigen binding. The power of VDJ recombination and N nucleotide addition has led to the common conception that the sequence of CDR-H3 is unrestricted in its variability and random in its composition. Under this view, the immune response is solely controlled by somatic positive and negative clonal selection mechanisms that act on individual B cells to promote production of protective antibodies and prevent the production of self-reactive antibodies. This concept of a repertoire of random antigen binding sites is inconsistent with the observation that diversity (D_H) gene segment sequence content by reading frame (RF) is evolutionarily conserved, creating biases in the prevalence and distribution of individual amino acids in CDR-H3. For example, arginine, which is often found in the CDR-H3 of dsDNA binding autoantibodies, is under-represented in the commonly used D_H RFs rearranged by deletion, but is a frequent component of rarely used inverted RF1 (iRF1), which is rearranged by inversion. To determine the effect of altering this germline bias in D_H gene segment sequence on autoantibody production, we generated mice that by genetic manipulation are forced to utilize an iRF1 sequence encoding two arginines. Over a one year period we collected serial serum samples from these unimmunized, specific pathogen-free mice and found that more than one-fifth of them contained elevated levels of dsDNA-binding IgG, but not IgM; whereas mice with a wild type D_H sequence did not. Thus, germline bias against the use of arginine enriched D_H sequence helps to reduce the likelihood of producing self-reactive antibodies.     

Metabolic Interplay between the Asian Citrus Psyllid and Its Profftella Symbiont: An Achilles’ Heel of the Citrus Greening Insect Vector

‘Candidatus Liberibacter asiaticus’ (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid. Interactions among D. citri and its microbial endosymbionts, including ‘Candidatus Profftella armatura’, are likely to impact transmission of CLas. We used quantitative mass spectrometry to compare the proteomes of CLas(+) and CLas(-) populations of D. citri, and found that proteins involved in polyketide biosynthesis by the endosymbiont Profftella were up-regulated in CLas(+) insects. Mass spectrometry analysis of the Profftella polyketide diaphorin in D. citri metabolite extracts revealed the presence of a novel diaphorin-related polyketide and the ratio of these two polyketides was changed in CLas(+) insects. Insect proteins differentially expressed between CLas(+) and CLas(-) D. citri included defense and immunity proteins, proteins involved in energy storage and utilization, and proteins involved in endocytosis, cellular adhesion, and cytoskeletal remodeling which are associated with microbial invasion of host cells. Insight into the metabolic interdependence between the insect vector, its endosymbionts, and the citrus greening pathogen reveals novel opportunities for control of this disease, which is currently having a devastating impact on citrus production worldwide.     

Limited catching bias in a wild population of birds with near‐complete census information

Animal research often relies on catching wild animals; however, individuals may have different trappability, and this can generate bias. We studied bias in mist netting, the main method for catching wild birds. The unusually high resighting rate in our study population—house sparrows (Passer domesticus) on Lundy Island (England)—allowed us to obtain accurate estimates of the population size. This unique situation enabled us to test for catching bias in mist netting using deviations from the expected Poisson distribution. There was no evidence that a fraction of the birds in the population consistently remained uncaught. However, we detected a different bias: More birds than expected were captured only once within a year. This bias probably resulted from a mixture of fieldworkers sometimes ignoring rapid recaptures and birds becoming net shy after their first capture. We had sufficient statistical power with the available data to detect a substantial uncaught fraction. Therefore, our data are probably unbiased toward catching specific individuals from our population. Our analyses demonstrate that intensively monitored natural insular populations, in which population size can be estimated precisely, provide the potential to address important unanswered questions without concerns about a fraction of the population remaining uncaught. Our approach can help researchers to test for catching bias in closely monitored wild populations for which reliable estimates of population size and dispersal are available.     

Serum tryptase detected during acute coronary syndrome is significantly related to the development of major adverse cardiovascular events after 2?years

Background

One of the greatest challenges in cardiovascular medicine is to define the best tools for performing an accurate risk stratification for the recurrence of ischemic events in acute coronary syndrome (ACS) patients.

Methods

We followed 65 ACS patients enrolled in a previous pilot study for 2 years after being discharged, focusing on the occurrence of major adverse cardiovascular events (MACE).The relationship between serum tryptase levels on admission, SYNergy between percutaneous coronary intervention with the TAXUS drug-eluting stent and the cardiac surgery score (SX-score), cardiovascular complexity and MACE at 2 years follow-up were analyzed.

Results

The ACS population was divided in two groups: patients with MACE (n = 23) and patients without MACE (n = 42).The tryptase measurement at admission (T0) and at discharge (T3) and SX-score were higher in patients who experienced MACE than in those without (p = 0.0001, p < 0.0001 and p = 0.006, respectively). Conversely, we found no significant association between MACE and C-reactive protein (CRP), and between MACE and maximum level of high-sensitivity troponin (hs-Tn) values.Among all patients with MACE, 96% belonged to the group that presented with cardiovascular complexity at the beginning of ACS index admission (p < 0.0001).The predictive accuracy of serum tryptase for MACE at follow up set at the cut-off point of 4.95 ng/ml at T0 and of 5.2 ng/ml at T3. Interestingly, patients with both the above cut-off tryptase values at T0 and at T3 presented a 1320% increase in the odds of developing MACE (p < 0.0001).

Conclusion

In ACS patients, serum tryptase measured during index admission is significantly correlated to the development of MACE up to 2 years, demonstrating a possible long-term prognostic role of this biomarker.

Electronic supplementary material

The online version of this article (doi:10.1186/s12948-015-0013-0) contains supplementary material, which is available to authorized users.