The lipid peroxidation metabolite 4-oxo-2-nonenal cross-links α-synuclein causing rapid formation of stable oligomers

Recently, the aldehyde 4-oxo-2-nonenal (ONE) was identified as a product of lipid peroxidation and found to be an effective protein modifier. In this in vitro study we investigated structural implications of the interaction between ONE and α-synuclein, a protein which forms intraneuronal inclusions in neurodegenerative disorders such as Parkinson’s disease and dementia with Lewy bodies. Our results demonstrate that ONE induced an almost complete conversion of monomeric α-synuclein into 40-80 nm wide and 6-8 nm high soluble β-sheet-rich oligomers with a molecular weight of ∼2000 kDa. Furthermore, the ONE-induced α-synuclein oligomers displayed a high stability and were not sensitive to treatment with sodium dodecyl sulfate, indicating that ONE stabilized the oligomers by cross-linking individual α-synuclein molecules. Despite prolonged incubation the oligomers did not continue to aggregate into a fibrillar state, thus suggesting that these α-synuclein species were not on a fibrillogenic pathway.     

The Novel α4B Murine α4 Integrin Protein Splicing Variant Inhibits α4 Protein-dependent Cell Adhesion

Integrins affect the motility of multiple cell types to control cell survival, growth, or differentiation, which are mediated by cell-cell and cell-extracellular matrix interactions. We reported previously that the α9 integrin splicing variant, SFα9, promotes WT α9 integrin-dependent adhesion. In this study, we introduced a new murine α4 integrin splicing variant, α4B, which has a novel short cytoplasmic tail. In inflamed tissues, the expression of α4B, as well as WT α4 integrin, was up-regulated. Cells expressing α4B specifically bound to VCAM-1 but not other α4 integrin ligands, such as fibronectin CS1 or osteopontin. The binding of cells expressing WT α4 integrin to α4 integrin ligands is inhibited by coexpression of α4B. Knockdown of α4B in metastatic melanoma cell lines results in a significant increase in lung metastasis. Expression levels of WT α4 integrin are unaltered by α4B, with α4B acting as a regulatory subunit for WT α4 integrin by a dominant-negative effect or inhibiting α4 integrin activation.     

Proteome analysis of whole saliva: a new tool for rheumatic diseases--the example of Sjögren's syndrome

Sj?gren's syndrome (pSS) is a systemic disease that affects salivary glands directly, and is therefore expected to influence the composition of human whole saliva (WS) fluid. The aim of this study was to characterize the WS proteins of pSS patients using a proteomic approach to assess a valid procedure to examine the global changes of the salivary protein profiles in connective tissue disorders. The WS proteins expressed in patients affected by pSS and healthy volunteers were analyzed using the 2-DE technique. The WS protein pattern was altered in pSS patients compared to controls, with a decrease in some of the typical salivary proteins. Particularly, a remarkable alteration of carbonic anhydrase VI was observed. Moreover, a comparison of WS protein profile of pSS patients with the one obtained from controls revealed a set of differentially expressed proteins. These proteins were related to acute and chronic inflammation while some others were involved in oxidative stress injury. These findings are in line with the systemic immuno-inflammatory aspects of pSS and open the possibility for a systematic search of diagnostic biomarkers and targets for therapeutic intervention in pSS.     

The membrane scaffold CD82 regulates cell adhesion by altering α4 integrin stability and molecular density

Hematopoietic stem/progenitor cell (HSPC) interactions with the bone marrow microenvironment are important for maintaining HSPC self-renewal and differentiation. In recent work, we identified the tetraspanin protein, CD82, as a regulator of HPSC adhesion and homing to the bone marrow, although the mechanism by which CD82 mediated adhesion was unclear. In the present study, we determine that CD82 expression alters cell–matrix adhesion, as well as integrin surface expression. By combining the superresolution microscopy imaging technique, direct stochastic optical reconstruction microscopy, with protein clustering algorithms, we identify a critical role for CD82 in regulating the membrane organization of α4 integrin subunits. Our data demonstrate that CD82 overexpression increases the molecular density of α4 within membrane clusters, thereby increasing cellular adhesion. Furthermore, we find that the tight packing of α4 into membrane clusters depend on CD82 palmitoylation and the presence of α4 integrin ligands. In combination, these results provide unique quantifiable evidence of CD82’s contribution to the spatial arrangement of integrins within the plasma membrane and suggest that regulation of integrin density by tetraspanins is a critical component of cell adhesion.     

The incorporation of the A2 protein to produce novel Qβ virus-like particles using cell-free protein synthesis

Virus-like particles (VLPs) have been employed for a number of nanometric applications because they self-assemble, exhibit a high degree of symmetry, and can be genetically and chemically modified. However, high symmetry does not allow for a single unique modification site on the VLP. Here, we demonstrate the co-expression of the cytotoxic A2 protein and the coat protein of the bacteriophage Qβ to form a nearly monodispersed population of novel VLPs. Cell-free protein synthesis allows for direct access and optimization of protein-synthesis and VLP-assembly. The A2 is shown to be incorporated at high efficiency, approaching a theoretical maximum of one A2 per VLP. This work demonstrates de novo production of a novel VLP, which contains a unique site that has the potential for future nanometric engineering applications.     

The study of iron mobilisation from transferrin using α-ketohydroxy heteroaromatic chelators

A group of heteroaromatic chelators with an α-ketohydroxy binding site have been tested for their ability to mobilise iron from transferrin in vitro. When these chelators were mixed with iron-saturated transferrin at physiological pH, biphasic reactions were observed. The α-ketohydroxy heteroaromatic chelators were found to cause substantial iron removal compared to other known chelators. These findings suggest that these chelators may have an important role in the study of iron metabolism and a possible clinical use in the treatment of transfusional iron overload in thalassaemia, and other diseases of iron imbalance.     

The clinical importance of CD4+CD7− in human diseases

The CD7 antigen is a member of the immunoglobulin superfamily that expresses on the surface of all thymocytes, a majority of mature T cells, and also natural killer cells. Interestingly, under physiological and different pathological conditions, the loss of CD7 antigen occurred in the subset of CD4⁺ memory T cells. Various functions have been proposed for CD7, including its role in the activation and intercellular adhesiveness of T cells. Several studies indicate that the number of CD4⁺CD7⁻ T cells increases in diseases such as chronic inflammation and T-cell malignancies, these being skin inflammatory lesions. Therefore, this can be useful for the diagnosis of cancer cells, especially with reference to blood origin, treatment monitoring, and establishment of new therapies. Therefore, a comprehensive review could be useful to increase our knowledge about the clinical importance of these cells in human disease.     

Differential Expression of Integrins α6 and α4 Determines Pathways in Human Peripheral CD4+ T Cell Differentiation

Integrins mediate leukocyte adhesion to vascular endothelium and thereby influence leukocyte recirculation. We have explored expression by peripheral blood T cells of β1 and β7 integrins, particularly α4β1 (VLA-4, CD49d), α4β7 (LPAM-1) and α6β1 (VLA-6, CD49f). Integrin expression differs between CD4+ cells and CD8+ cells in that CD4+ cells: 1) are more heterogeneous, particularly for α4; 2) express on the average less α4 and β7; and 3) express on the average more α6 and β1.2D gel electrophoretic analysis was combined with flow cytometric analysis to determine which integrin chain pairs are expressed by the CD45RO – (naive) and CD45RO+ (memory) subsets of CD4+ cells. CD45RO– (naive) cells express homogeneously at intermediate levels the three integrin pairs α6β1, α4β1 and α4β7. Although 2D gel analysis suggests similar average integrin chain composition for CD45RO+CD4+ (memory) cells, flow cytometric analysis demonstrates multiple subsets of CD45RO+ cells differing markedly from each other and from naive cells in levels of expression of α6 and α4 integrins. There are a minimum of three CD45RO+ subsets: 1) α4β1^hiα6β1^hiα4β7^neg which comprises the majority of memory cells; 2) α4β7^hiα6β1^low presumptive gut-homing memory cells; and 3) α6β1^hiα4β7^negα4β1^neg, a previously unidentified subset expected to have unique migrational-functional properties. Of particular importance in these results are: the expression by CD4+ naive cells of α6β1, α4β1 and α4β7, the overall prominence and regulation of α6β1 on CD4+ cells, and the selective decreases as well as increases in α4β7 and α4β1 during CD4+ memory specialization. Taken together, these results suggest that differential regulation of expression of α4 and α6 integrin chains that accompany naive-to-memory transition in CD4+ cells are instrumental in generating functional subsets of CD4+ memory cells with specialized recirculation abilities.     

The effects of whole body vibration on humans: dangerous or advantageous?

The effects of whole body vibration (WBV) have been studied extensively in occupational medicine. In particular, it has been shown that when the body undergoes chronically to whole body vibrations spinal degeneration is likely to be one of the deleterious outcomes. Low back pain has been shown to be the leading major cause of industrial disability in the population under the age of 45 years and has been linked to whole body vibration exposure encountered in some industrial settings. Whole body vibration has been recently purposed as an exercise intervention suggesting its effectiveness in increasing force-generating capacity in lower limbs and low back. It has also been reported to be an effective non-pharmacological intervention for patients with low back pain. Relatively short exposure to whole body vibration has been also shown to increase the serum levels of testosterone and growth hormone. The combined effects on the neuromuscular system and endocrine system seem to suggest its effectiveness as a therapeutic approach for sarcopenia and possibly osteoporosis. Due to the danger of long-term exposure to whole body vibration, it is important to develop safe exercise protocols in order to determine exercise programs for different populations.     

The α-Actinin Gene Family: A Revised Classification

The sequencing of a genome is the first stage of its complete characterization. Subsequent work seeks to utilize available sequence data to gain a better understanding of the genes which are found within a genome. Gene families comprise large portions of the genomes of higher vertebrates, and the available genomic data allow for a reappraisal of gene family evolution. This reappraisal will clarify relatedness within and between gene families. One such family, the alpha-actinin gene family, is part of the spectrin superfamily. There are four known loci, which encode alpha-actinins 1, 2, 3, and 4. Of the eight domains in alpha-actinin, the actin-binding domain is the most highly conserved. Here we present evidence gained through phylogenetic analyses of the highly conserved actin-binding domain that alpha-actinin 2 was the first of the four alpha-actinins to arise by gene duplication, followed by the divergence of alpha-actinin 3 and then alpha-actinins 1 and 4. Resolution of the gene tree for this gene family has allowed us to reclassify several alpha-actinins which were previously given names inconsistent with the most widely accepted nomenclature for this gene family. This reclassification clarifies previous discrepancies in the public databases as well as in the literature, thus eliminating confusion caused by continued misclassification of members of the alpha-actinin gene family. In addition, the topology found for this gene family undermines the 2R hypothesis theory of two rounds of genome duplication early in vertebrate evolution.     

The α-Actinin Gene Family: A Revised Classification

Abstract The sequencing of a genome is the first stage of its complete characterization. Subsequent work seeks to utilize available sequence data to gain a better understanding of the genes which are found within a genome. Gene families comprise large portions of the genomes of higher vertebrates, and the available genomic data allow for a reappraisal of gene family evolution. This reappraisal will clarify relatedness within and between gene families. One such family, the α-actinin gene family, is part of the spectrin superfamily. There are four known loci, which encode α-actinins 1, 2, 3, and 4. Of the eight domains in α-actinin, the actin-binding domain is the most highly conserved. Here we present evidence gained through phylogenetic analyses of the highly conserved actin-binding domain that α-actinin 2 was the first of the four α-actinins to arise by gene duplication, followed by the divergence of α-actinin 3 and then α-actinins 1 and 4. Resolution of the gene tree for this gene family has allowed us to reclassify several α-actinins which were previously given names inconsistent with the most widely accepted nomenclature for this gene family. This reclassification clarifies previous discrepancies in the public databases as well as in the literature, thus eliminating confusion caused by continued misclassification of members of the α-actinin gene family. In addition, the topology found for this gene family undermines the 2R hypothesis theory of two rounds of genome duplication early in vertebrate evolution.     

The effects of α-synuclein on phospholipid vesicle integrity: a study using 31P NMR and electron microscopy

Associations between the 140 amino acid protein α-synuclein (asyn) and presynaptic vesicles may play a role in maintaining synaptic plasticity and neurotransmitter release. These physiological processes may involve disruption and fusion of vesicles, arising from interactions between specific regions of asyn, including the highly basic N-terminal domain, and the surface of vesicles. This work investigates whether asyn affects the integrity of model unilamellar vesicles of varying size and phospholipid composition, by monitoring paramagnetic Mn²⁺-induced broadening of peaks in the ³¹P nuclear magnetic resonance spectrum of the lipid head groups. It is shown that asyn increases the permeability to Mn²⁺ of both large (200nm diameter) and small (50nm diameter) vesicles composed of zwitterionic phosphatidylcholine and anionic phosphatidylglycerol at protein/lipid molar ratios as low as 1:2000. Further experiments on peptides corresponding to sequences in the N-terminal (10–48), C-terminal (120–140) and central hydrophobic (71–82) regions of asyn suggest that single regions of the protein are capable of permeabilizing the vesicles to varying extents. Electron micrographs of the vesicles after addition of asyn indicate that the enhanced permeability is coupled to large-scale disruption or fusion of the vesicles. These results indicate that asyn is able to permeabilize phospholipid vesicles at low relative concentrations, dependent upon the properties of the vesicles. This could have implications for asyn playing a role in vesicle synthesis, maintenance and fusion within synapses.     

The disappearing CD4(+)T cells in HIV infection: a case of over-stimulation?

Infection with the human immunodeficiency virus (HIV) causes a gradual decline in essential immune-system cells called CD4(+)"helper" T cells. These cells are also principal viral targets, but, paradoxically, direct cell-killing does not explain their disappearance. HIV also induces a chronic and increasing state of immune activation. In a mathematical model of normal T-cell kinetics incorporating a cytokine growth factor, increased activation alone explains these T-cell losses, a switch from "na?ve" to "memory" phenotype, and certain other features of HIV disease.