Binding interface change and cryptic variation in the evolution of protein-protein interactions

Background:Physical interactions between proteins are essential for almost all biological functions and systems. To understand the evolution of function it is therefore important to understand the evolution of molecular interactions. Of key importance is the evolution of binding specificity, the set of interactions made by a protein, since change in specificity can lead to “rewiring” of interaction networks. Unfortunately, the interfaces through which proteins interact are complex, typically containing many amino-acid residues that collectively must contribute to binding specificity as well as binding affinity, structural integrity of the interface and solubility in the unbound state. Results: In order to study the relationship between interface composition and binding specificity, we make use of paralogous pairs of yeast proteins. Immediately after duplication these paralogues will have identical sequences and protein products that make an identical set of interactions. As the sequences diverge, we can correlate amino-acid change in the interface with any change in the specificity of binding. We show that change in interface regions correlates only weakly with change in specificity, and many variants in interfaces are functionally equivalent. We show that many of the residue replacements within interfaces are silent with respect to their contribution to binding specificity. Conclusions: We conclude that such functionally-equivalent change has the potential to contribute to evolutionary plasticity in interfaces by creating cryptic variation, which in turn may provide the raw material for functional innovation and coevolution.BBSRCWellcome Trust Institutional Strategic Support Awar

Pivotal Role of Inosine Triphosphate Pyrophosphatase in Maintaining Genome Stability and the Prevention of Apoptosis in Human Cells

Pure nucleotide precursor pools are a prerequisite for high-fidelity DNA replication and the suppression of mutagenesis and carcinogenesis. ITPases are nucleoside triphosphate pyrophosphatases that clean the precursor pools of the non-canonical triphosphates of inosine and xanthine. The precise role of the human ITPase, encoded by the ITPA gene, is not clearly defined. ITPA is clinically important because a widespread polymorphism, 94C>A, leads to null ITPase activity in erythrocytes and is associated with an adverse reaction to thiopurine drugs. We studied the cellular function of ITPA in HeLa cells using the purine analog 6-N hydroxylaminopurine (HAP), whose triphosphate is also a substrate for ITPA. In this study, we demonstrate that ITPA knockdown sensitizes HeLa cells to HAP-induced DNA breaks and apoptosis. The HAP-induced DNA damage and cytotoxicity observed in ITPA knockdown cells are rescued by an overexpression of the yeast ITPase encoded by the HAM1 gene. We further show that ITPA knockdown results in elevated mutagenesis in response to HAP treatment. Our studies reveal the significance of ITPA in preventing base analog-induced apoptosis, DNA damage and mutagenesis in human cells. This implies that individuals with defective ITPase are predisposed to genome damage by impurities in nucleotide pools, which is drastically augmented by therapy with purine analogs. They are also at an elevated risk for degenerative diseases and cancer

Diet supplementation for 5 weeks with polyphenol-rich cereals improves several functions and the redox state of mouse leucocytes

BACKGROUND: Cereals naturally contain a great variety of polyphenols, which exert a wide range of physiological effects both in vitro and in vivo. Many of their protective effects, including an improvement of the function and redox state of immune cells in unhealthy or aged subjects come from their properties as powerful antioxidant compounds. However, whether cereal-based dietary supplementation positively affects the immune function and cellular redox state of healthy subjects remains unclear. AIM OF THE STUDY: To investigate the effects of supplementation (20% wt/wt) for 5 weeks with four different cereal fractions on healthy mice. METHODS: Several parameters of function and redox state of peritoneal leukocytes were measured. The cereals, named B (wheat germ), C (buckwheat flour), D (fine rice bran) and E (wheat middlings) contained different amounts of gallic acid, p-hydroxybenzoic acid, vanillic acid, sinapic acid, p-coumaric acid, ferulic acid, quercetin, catechin, rutin and oryzanol as major polyphenols. RESULTS: In general, all cereal fractions caused an improvement of the leukocyte parameters studied such as chemotaxis capacity, microbicidal activity, lymphoproliferative response to mitogens, interleukin-2 (IL-2) and tumor necrosis factor (TNFα) release, as well as oxidized glutathione (GSSG), GSSG/GSH ratio, catalase (CAT) activity and lipid oxidative damage. We observed similar effects among the cereal fractions. CONCLUSIONS: The results suggest that some of these effects may due, at least partially, to the antioxidant activity of the polyphenols naturally present in cereals. Since an appropriate function of the leukocytes has been proposed as marker of the health state, a short-term intake of cereals seems to be sufficient to exert a benefit in the health of the general population. However, further studies are needed to assess the optimal doses and to find out which active polyphenols are able to mediate the observed physiological effects before recommending their regular consumption

Complexity absorption : A processual strategic approach to corporate entrepreneurship strategy

The author Haina Zhang acknowledges the support of research grant received from the Lancaster University Research Committee (award reference: SGS21/33). The author is also grateful to Professor David Brown for his suggestions on earlier version of this paper.Peer reviewedPublisher PD

Cued to act on impulse: more impulsive choice and risky decision making by women susceptible to overeating after exposure to food stimuli

There is increasing evidence that individual differences in tendency to overeat relate to impulsivity, possibly by increasing reactivity to food-related cues in the environment. This study tested whether acute exposure to food cues enhanced impulsive and risky responses in women classified on tendency to overeat, indexed by scores on the three factor eating questionnaire disinhibition (TFEQ-D), restraint (TFEQ-R) and hunger scales. Ninety six healthy women completed two measures of impulsive responding (delayed discounting, DDT and a Go No-Go, GNG, task) and a measure of risky decision making (the balloon analogue risk task, BART) as well as questionnairemeasures of impulsive behaviour either after looking at a series of pictures of food or visually matched controls. Impulsivity (DDT) and risk-taking (BART) were both positively associated with TFEQ-D scores, but in both cases this effect was exacerbated by prior exposure to food cues. No effects of restraint were found. TFEQ-D scores were also related tomore commission errors on the GNG, while restrained women were slower on the GNG, but neither effect was modified by cue exposure. Overall these data suggest that exposure to food cues act to enhance general impulsive responding in women at risk of overeating and tentatively suggest an important interaction between tendency for impulsive decision making and food cues thatmay help explain a key underlying risk factor for overeating

Three dimensional electron microscopy reveals changing axonal and myelin morphology along normal and partially injured optic nerves

Following injury to the central nervous system, axons and myelin distinct from the initial injury site undergo changes associated with compromised function. Quantifying such changes is important to understanding the pathophysiology of neurotrauma; however, most studies to date used 2 dimensional (D) electron microscopy to analyse single sections, thereby failing to capture changes along individual axons. We used serial block face scanning electron microscopy (SBF SEM) to undertake 3D reconstruction of axons and myelin, analysing optic nerves from normal uninjured female rats and following partial optic nerve transection. Measures of axon and myelin dimensions were generated by examining 2D images at 5 µm intervals along the 100 µm segments. In both normal and injured animals, changes in axonal diameter, myelin thickness, fiber diameter, G-ratio and percentage myelin decompaction were apparent along the lengths of axons to varying degrees. The range of values for axon diameter along individual reconstructed axons in 3D was similar to the range from 2D datasets, encompassing reported variation in axonal diameter attributed to retinal ganglion cell diversity. 3D electron microscopy analyses have provided the means to demonstrate substantial variability in ultrastructure along the length of individual axons and to improve understanding of the pathophysiology of neurotrauma

Acute reduction of serum 8-iso-PGF2-alpha and advanced oxidation protein products in vivo by a polyphenol-rich beverage; a pilot clinical study with phytochemical and in vitro antioxidant characterization

<p>Abstract</p> <p>Background</p> <p>Measuring the effects of the acute intake of natural products on human biomarker concentrations, such as those related to oxidation and inflammation, can be an advantageous strategy for early clinical research on an ingredient or product.</p> <p>Methods</p> <p>31 total healthy subjects were randomized in a double-blinded, placebo-controlled, acute pilot study with post-hoc subgroup analysis on 20 of the subjects. The study examined the effects of a single dose of a polyphenol-rich beverage (PRB), commercially marketed as "SoZo^®", on serum anti-inflammatory and antioxidant markers. In addition, phytochemical analyses of PRB, and <it>in vitro </it>antioxidant capacity were also performed.</p> <p>Results</p> <p>At 1 hour post-intake, serum values for 8-iso-PGF2-alpha and advanced oxidation protein products decreased significantly by 40% and 39%, respectively. Additionally, there was a trend toward decreased C-reactive protein, and increased nitric oxide levels. Both placebo and PRB treatment resulted in statistically significant increases in hydroxyl radical antioxidant capacity (HORAC) compared to baseline; PRB showed a higher percent change (55-75% versus 23-74% in placebo group), but the two groups did not differ significantly from each other.</p> <p>Conclusions</p> <p>PRB produced statistically significant changes in several blood biomarkers related to antioxidant/anti-inflammatory effects. Future studies are justified to verify results and test for cumulative effects of repeated intakes of PRB. The study demonstrates the potential utility of acute biomarker measurements for evaluating antioxidant/anti-inflammatory effects of natural products.</p

Identification of Candida glabrata genes involved in pH modulation and modification of the phagosomal environment in macrophages

notes: PMCID: PMC4006850types: Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov'tCandida glabrata currently ranks as the second most frequent cause of invasive candidiasis. Our previous work has shown that C. glabrata is adapted to intracellular survival in macrophages and replicates within non-acidified late endosomal-stage phagosomes. In contrast, heat killed yeasts are found in acidified matured phagosomes. In the present study, we aimed at elucidating the processes leading to inhibition of phagosome acidification and maturation. We show that phagosomes containing viable C. glabrata cells do not fuse with pre-labeled lysosomes and possess low phagosomal hydrolase activity. Inhibition of acidification occurs independent of macrophage type (human/murine), differentiation (M1-/M2-type) or activation status (vitamin D3 stimulation). We observed no differential activation of macrophage MAPK or NFκB signaling cascades downstream of pattern recognition receptors after internalization of viable compared to heat killed yeasts, but Syk activation decayed faster in macrophages containing viable yeasts. Thus, delivery of viable yeasts to non-matured phagosomes is likely not triggered by initial recognition events via MAPK or NFκB signaling, but Syk activation may be involved. Although V-ATPase is abundant in C. glabrata phagosomes, the influence of this proton pump on intracellular survival is low since blocking V-ATPase activity with bafilomycin A1 has no influence on fungal viability. Active pH modulation is one possible fungal strategy to change phagosome pH. In fact, C. glabrata is able to alkalinize its extracellular environment, when growing on amino acids as the sole carbon source in vitro. By screening a C. glabrata mutant library we identified genes important for environmental alkalinization that were further tested for their impact on phagosome pH. We found that the lack of fungal mannosyltransferases resulted in severely reduced alkalinization in vitro and in the delivery of C. glabrata to acidified phagosomes. Therefore, protein mannosylation may play a key role in alterations of phagosomal properties caused by C. glabrata.Deutsche ForschungsgemeinschaftNational Institutes for HealthWellcome TrustBBSR