Assessing the utility of conserving evolutionary history

It is often claimed that conserving evolutionary history is more efficient than species‐based approaches for capturing the attributes of biodiversity that benefit people. This claim underpins academic analyses and recommendations about the distribution and prioritization of species and areas for conservation, but evolutionary history is rarely considered in practical conservation activities. One impediment to implementation is that arguments related to the human‐centric benefits of evolutionary history are often vague and the underlying mechanisms poorly explored. Herein we identify the arguments linking the prioritization of evolutionary history with benefits to people, and for each we explicate the purported mechanism, and evaluate its theoretical and empirical support. We find that, even after 25 years of academic research, the strength of evidence linking evolutionary history to human benefits is still fragile. Most – but not all – arguments rely on the assumption that evolutionary history is a useful surrogate for phenotypic diversity. This surrogacy relationship in turn underlies additional arguments, particularly that, by capturing more phenotypic diversity, evolutionary history will preserve greater ecosystem functioning, capture more of the natural variety that humans prefer, and allow the maintenance of future benefits to humans. A surrogate relationship between evolutionary history and phenotypic diversity appears reasonable given theoretical and empirical results, but the strength of this relationship varies greatly. To the extent that evolutionary history captures unmeasured phenotypic diversity, maximizing the representation of evolutionary history should capture variation in species characteristics that are otherwise unknown, supporting some of the existing arguments. However, there is great variation in the strength and availability of evidence for benefits associated with protecting phenotypic diversity. There are many studies finding positive biodiversity–ecosystem functioning relationships, but little work exists on the maintenance of future benefits or the degree to which humans prefer sets of species with high phenotypic diversity or evolutionary history. Although several arguments link the protection of evolutionary history directly with the reduction of extinction rates, and with the production of relatively greater future biodiversity via increased adaptation or diversification, there are few direct tests. Several of these putative benefits have mismatches between the relevant spatial scales for conservation actions and the spatial scales at which benefits to humans are realized. It will be important for future work to fill in some of these gaps through direct tests of the arguments we define here.     

Characterization of glycosomal RING finger proteins of trypanosomatids

The glycosomes of trypanosomatids are essential organelles that are evolutionarily related to peroxisomes of other eukaryotes. The peroxisomal RING proteins-PEX2, PEX10 and PEX12-comprise a network of integral membrane proteins that function in the matrix protein import cycle. Here, we describe PEX10 and PEX12 in Trypanosoma brucei, Leishmania major, and Trypanosoma cruzi. We expressed GFP fusions of each T. brucei coding region in procyclic form T. brucei, where they localized to glycosomes and behaved as integral membrane proteins. Despite the weak transmembrane predictions for TbPEX12, protease protection assays demonstrated that both the N and C termini are cytosolic, similar to mammalian PEX12. GFP fusions of T. cruzi PEX10 and L. major PEX12 also localized to glycosomes in T. brucei indicating that glycosomal membrane protein targeting is conserved across trypanosomatids.     

Acceptor specificity of the Pasteurella hyaluronan and chondroitin synthases and production of chimeric glycosaminoglycans

The hyaluronan (HA) synthase, PmHAS, and the chondroitin synthase, PmCS, from the Gram-negative bacterium Pasteurella multocida polymerize the glycosaminoglycan (GAG) sugar chains HA or chondroitin, respectively. The recombinant Escherichia coli-derived enzymes were shown previously to elongate exogenously supplied oligosaccharides of their cognate GAG (e.g. HA elongated by PmHAS). Here we show that oligosaccharides and polysaccharides of certain noncognate GAGs (including sulfated and iduronic acid-containing forms) are elongated by PmHAS (e.g. chondroitin elongated by PmHAS) or PmCS. Various acceptors were tested in assays where the synthase extended the molecule with either a single monosaccharide or a long chain (approximately 10(2-4) sugars). Certain GAGs were very poor acceptors in comparison to the cognate molecules, but elongated products were detected nonetheless. Overall, these findings suggest that for the interaction between the acceptor and the enzyme (a) the orientation of the hydroxyl at the C-4 position of the hexosamine is not critical, (b) the conformation of C-5 of the hexuronic acid (glucuronic versus iduronic) is not crucial, and (c) additional negative sulfate groups are well tolerated in certain cases, such as on C-6 of the hexosamine, but others, including C-4 sulfates, were not or were poorly tolerated. In vivo, the bacterial enzymes only process unsulfated polymers; thus it is not expected that the PmCS and PmHAS catalysts would exhibit such relative relaxed sugar specificity by acting on a variety of animal-derived sulfated or epimerized GAGs. However, this feature allows the chemoenzymatic synthesis of a variety of chimeric GAG polymers, including mimics of proteoglycan complexes.     

D1 and D2 dopamine receptor expression is regulated by direct interaction with the chaperone protein calnexin

As for all proteins, G protein-coupled receptors (GPCRs) undergo synthesis and maturation within the endoplasmic reticulum (ER). The mechanisms involved in the biogenesis and trafficking of GPCRs from the ER to the cell surface are poorly understood, but they may involve interactions with other proteins. We have now identified the ER chaperone protein calnexin as an interacting protein for both D(1) and D(2) dopamine receptors. These protein-protein interactions were confirmed using Western blot analysis and co-immunoprecipitation experiments. To determine the influence of calnexin on receptor expression, we conducted assays in HEK293T cells using a variety of calnexin-modifying conditions. Inhibition of glycosylation either through receptor mutations or treatments with glycosylation inhibitors partially blocks the interactions with calnexin with a resulting decrease in cell surface receptor expression. Confocal fluorescence microscopy reveals the accumulation of D(1)-green fluorescent protein and D(2)-yellow fluorescent protein receptors within internal stores following treatment with calnexin inhibitors. Overexpression of calnexin also results in a marked decrease in both D(1) and D(2) receptor expression. This is likely because of an increase in ER retention because confocal microscopy revealed intracellular clustering of dopamine receptors that were co-localized with an ER marker protein. Additionally, we show that calnexin interacts with the receptors via two distinct mechanisms, glycan-dependent and glycan-independent, which may underlie the multiple effects (ER retention and surface trafficking) of calnexin on receptor expression. Our data suggest that optimal receptor-calnexin interactions critically regulate D(1) and D(2) receptor trafficking and expression at the cell surface, a mechanism likely to be of importance for many GPCRs.     

Tau phosphorylation by GSK-3β promotes tangle-like filament morphology

Background

Neurofibrillary tangles (NFTs) are intraneuronal aggregates associated with several neurodegenerative diseases including Alzheimer's disease. These abnormal accumulations are primarily comprised of fibrils of the microtubule-associated protein tau. During the progression of NFT formation, disperse and non-interacting tau fibrils become stable aggregates of tightly packed and intertwined filaments. Although the molecular mechanisms responsible for the conversion of disperse tau filaments into tangles of filaments are not known, it is believed that some of the associated changes in tau observed in Alzheimer's disease, such as phosphorylation, truncation, ubiquitination, glycosylation or nitration, may play a role.

Results

We have investigated the effects of tau phosphorylation by glycogen synthase kinase-3β (GSK-3β) on tau filaments in an in vitro model system. We have found that phosphorylation by GSK-3β is sufficient to cause tau filaments to coalesce into tangle-like aggregates similar to those isolated from Alzheimer's disease brain.

Conclusion

These results suggest that phosphorylation of tau by GSK-3β promotes formation of tangle-like filament morphology. The in vitro cell-free experiments described here provide a new model system to study mechanisms of NFT development. Although the severity of dementia has been found to correlate with the presence of NFTs, there is some question as to the identity of the neurotoxic agents involved. This model system will be beneficial in identifying intermediates or side reaction products that might be neurotoxic.     

An exploratory study of cardiac function and oxygen uptake during cycle ergometry in overweight children

Objective: Obesity has been proposed to negatively impact cardiac function in overweight (OW) individuals. The relationship between diastolic dysfunction and oxygen uptake (V?o ₂) kinetics is equivocal. This exploratory investigation evaluated the relationship between resting left ventricular function and V?o ₂ kinetics during cycle ergometry in OW and non‐overweight (NO) children and adolescents. Research Methods and Procedures: Fourteen OW (>85 percentile for BMI for age and gender) children, 10 boys and 4 girls (age, 11.7 ± 1.9 years; body mass, 80.6 ± 45.5 kg) and 10 NO children (4 boys, 6 girls) volunteered to participate in the study (age, 12.5 ± 2.1 years; body mass, 45.8 ± 13.8 kg). Resting cardiovascular structure and function were assessed using spectral Doppler echocardiography. All subjects underwent two sub‐maximal exercise stages on a cycle ergometer (3 minutes unloaded and 5 minutes at 50 W, both at a cadence of 50 rpm). Respiratory data were measured on a breath‐by‐breath basis at both workloads and the mean response time (MRT) was calculated. Results: Analysis of the MRT data demonstrated that there were no significant differences between OW and NO (OW, 52.6 ± 11.7 seconds vs. NO, 45.6 ± 7.4 seconds). Significant correlations (p < 0.05) were obtained between MRT V?o ₂ and echocardiographic‐derived mitral valve inflow pressure half‐time (r = 0.55) and between MRT V?o ₂, and mitral valve inflow deceleration time (r = 0.55). Discussion: The evidence from this research suggests a possible link between left ventricular diastolic function at rest and oxygen uptake kinetics during sub‐maximal exercise in OW and NO children and adolescents.     

Identification of novel high-frequency DNA methylation changes in breast cancer

Recent data have revealed that epigenetic alterations, including DNA methylation and chromatin structure changes, are among the earliest molecular abnormalities to occur during tumorigenesis. The inherent thermodynamic stability of cytosine methylation and the apparent high specificity of the alterations for disease may accelerate the development of powerful molecular diagnostics for cancer. We report a genome-wide analysis of DNA methylation alterations in breast cancer. The approach efficiently identified a large collection of novel differentially DNA methylated loci (approximately 200), a subset of which was independently validated across a panel of over 230 clinical samples. The differential cytosine methylation events were independent of patient age, tumor stage, estrogen receptor status or family history of breast cancer. The power of the global approach for discovery is underscored by the identification of a single differentially methylated locus, associated with the GHSR gene, capable of distinguishing infiltrating ductal breast carcinoma from normal and benign breast tissues with a sensitivity and specificity of 90% and 96%, respectively. Notably, the frequency of these molecular abnormalities in breast tumors substantially exceeds the frequency of any other single genetic or epigenetic change reported to date. The discovery of over 50 novel DNA methylation-based biomarkers of breast cancer may provide new routes for development of DNA methylation-based diagnostics and prognostics, as well as reveal epigenetically regulated mechanism involved in breast tumorigenesis.