Alcoholism is a devastating brain disorder that affects millions of people worldwide. The development of alcoholism is caused by alcohol-induced maladaptive changes in neural circuits involved in emotions, motivation, and decision-making. Because of its involvement in these processes, the amygdala is thought to be a key neural structure involved in alcohol addiction. However, the molecular mechanisms that govern the development of alcoholism are incompletely understood. We have previously shown that in a limited access choice paradigm, C57BL/6J mice progressively escalate their alcohol intake and display important behavioral characteristic of alcohol addiction, in that they become insensitive to quinine-induced adulteration of alcohol. This study used the limited access choice paradigm to study gene expression changes in the amygdala during the escalation to high alcohol consumption in C57BL/6J mice. Microarray analysis revealed that changes in gene expression occurred predominantly after one week, i.e. during the initial escalation of alcohol intake. One gene that stood out from our analysis was the adapter protein 14-3-3ζ, which was up-regulated during the transition from low to high alcohol intake. Independent qPCR analysis confirmed the up-regulation of amygdala 14-3-3ζ during the escalation of alcohol intake. Subsequently, we found that local knockdown of 14-3-3ζ in the amygdala, using RNA interference, dramatically augmented alcohol intake. In addition, knockdown of amygdala 14-3-3ζ promoted the development of inflexible alcohol drinking, as apparent from insensitivity to quinine adulteration of alcohol. This study identifies amygdala 14-3-3ζ as a novel key modulator that is engaged during escalation of alcohol use.

 Open Access (OA) to scientific information is an important step forward in communication patterns, yet we still need to reinforce OA principles to promote a cultural change of traditional publishing practices. The advantages of free access to scientific information are even more evident in public health where knowledge is directly associated with human wellbeing.

Porcine reproductive and respiratory syndrome virus (PRRSV) is an arterivirus that shows a restricted in vivo tropism for subsets of porcine macrophages, with alveolar macrophages being major target cells. The virus is associated with respiratory problems in pigs of all ages and is commonly isolated on farms with porcine respiratory disease complex (PRDC). Due to virus-induced macrophage death early in infection, PRRSV hampers the innate defence against pathogens in the lungs. In addition, the virus might also directly affect the antimicrobial functions of macrophages. This study examined whether interaction of European genotype PRRSV with primary alveolar macrophages (PAM) affects their phagocytic capacity. Inoculation of macrophages with both subtype I PRRSV (LV) and subtype III PRRSV (Lena) showed that the virus inhibits PAM phagocytosis. Similar results were obtained using inactivated PRRSV (LV), showing that initial interaction of the virion with the cell is sufficient to reduce phagocytosis, and that no productive infection is required. When macrophages were incubated with sialoadhesin- (Sn) or CD163-specific antibodies, two entry mediators of the virus, only Sn-specific antibodies downregulated the phagocytic capacity of PAM, indicating that interaction with Sn, but not CD163, mediates the inhibitory effect of PRRSV on phagocytosis. In conclusion, this study shows that European genotype PRRSV inhibits PAM phagocytosis in vitro, through the interaction with its internalization receptor Sn. If similar events occur in vivo, this interaction may be important in the development of PRDC, as often seen in the field.

Diabetes mellitus is associated with micro- and macrovascular complications and increased cardiovascular risk. Elevated levels of serum asymmetric dimethylarginine (ADMA) may be responsible for endothelial dysfunction associated with diabetes-induced vascular impairment. Vitamin D may have potential protective effects against arterial stiffening. This study aimed to examine both the effects of diabetes on the functional/structural properties of the aorta and the endothelial function and the effects of vitamin D supplementation.

A whole-genome genotyping array has previously been developed for Malus using SNP data from 28 Malus genotypes. This array offers the prospect of high throughput genotyping and linkage map development for any given Malus progeny. To test the applicability of the array for mapping in diverse Malus genotypes, we applied the array to the construction of a SNP-based linkage map of an apple rootstock progeny.

This study examines the dynamic co-localization of lipid-anchored fluorescent proteins in living cells using pulsed-interleaved excitation fluorescence cross-correlation spectroscopy (PIE-FCCS) and fluorescence lifetime analysis. Specifically, we look at the pairwise co-localization of anchors from lymphocyte cell kinase (LCK: myristoyl, palmitoyl, palmitoyl), RhoA (geranylgeranyl), and K-Ras (farnesyl) proteins in different cell types. In Jurkat cells, a density-dependent increase in cross-correlation among RhoA anchors is observed, while LCK anchors exhibit a more moderate increase and broader distribution. No correlation was detected among K-Ras anchors or between any of the different anchor types studied. Fluorescence lifetime data reveal no significant Förster resonance energy transfer in any of the data. In COS 7 cells, minimal correlation was detected among LCK or RhoA anchors. Taken together, these observations suggest that some lipid anchors take part in anchor-specific co-clustering with other existing clusters of native proteins and lipids in the membrane. Importantly, these observations do not support a simple interpretation of lipid anchor-mediated organization driven by partitioning based on binary lipid phase separation.

Many Gram-positive and Gram-negative bacteria produce large quantities of indole as an intercellular signal in microbial communities. Indole demonstrated to affect gene expression in Escherichia coli as an intra-species signaling molecule. In contrast to E. coli, Salmonella does not produce indole because it does not harbor tnaA, which encodes the enzyme responsible for tryptophan metabolism. Our previous study demonstrated that E. coli-conditioned medium and indole induce expression of the AcrAB multidrug efflux pump in Salmonella enterica serovar Typhimurium for inter-species communication; however, the global effect of indole on genes in Salmonella remains unknown.

Breast cancer survivors, particularly those treated with chemotherapy, are at significantly increased risk for long-term cognitive and neurobiologic impairments. These deficits tend to involve skills that are subserved by distributed brain networks. Additionally, neuroimaging studies have shown a diffuse pattern of brain structure changes in chemotherapy-treated breast cancer survivors that might impact large-scale brain networks.

Ankylosing spondylitis (AS) is a familial, heritable disease specified by syndesmophyte formation leading to an ankylosed spine. Endoplasmic reticulum aminopeptidase 1 (ERAP1) genetic variations have been widely proved to be associated with AS in several ethnic populations. The aim of this study was to investigate whether ERAP1 single nucleotide polymorphisms (SNPs) are associated with AS susceptibility and disease severity in Taiwanese.

AMPA-type glutamate receptors (AMPARs) are responsible for a variety of processes in the mammalian brain including fast excitatory neurotransmission, postsynaptic plasticity, or synapse development. Here, with comprehensive and quantitative proteomic analyses, we demonstrate that native AMPARs are macromolecular complexes with a large molecular diversity. This diversity results from coassembly of the known AMPAR subunits, pore-forming GluA and three types of auxiliary proteins, with 21 additional constituents, mostly secreted proteins or transmembrane proteins of different classes. Their integration at distinct abundance and stability establishes the heteromultimeric architecture of native AMPAR complexes: a defined core with a variable periphery resulting in an apparent molecular mass between 0.6 and 1 MDa. The additional constituents change the gating properties of AMPARs and provide links to the protein dynamics fundamental for the complex role of AMPARs in formation and operation of glutamatergic synapses.

Akt, a serine-threonine protein kinase, exists as three isoforms. The Akt signaling pathway controls multiple cellular functions in the cardiovascular system, and the atheroprotective endothelial cell-dependent role of Akt1 has been recently demonstrated. The role of Akt3 isoform in cardiovascular pathophysiology is not known. We explored the role of Akt3 in atherosclerosis using mice with a genetic ablation of the Akt3 gene. Using hyperlipidemic ApoE(-/-) mice, we demonstrated a macrophage-dependent, atheroprotective role for Akt3. In vitro experiments demonstrated differential subcellular localization of Akt1 and Akt3 in macrophages and showed that Akt3 specifically inhibits macrophage cholesteryl ester accumulation and foam cell formation, a critical early event in atherogenesis. Mechanistically, Akt3 suppresses foam cell formation by reducing lipoprotein uptake and promoting ACAT-1 degradation via the ubiquitin-proteasome pathway. These studies demonstrate the nonredundant atheroprotective role for Akt3 exerted via the previously unknown link between the Akt signaling pathway and cholesterol metabolism.

AnkyrinG (ankG) is highly enriched in neurons at axon initial segments (AISs) where it clusters Na(+) and K(+) channels and maintains neuronal polarity. How ankG becomes concentrated at the AIS is unknown. Here, we show that as neurons break symmetry, they assemble a distal axonal submembranous cytoskeleton, comprised of ankyrinB (ankB), αII-spectrin, and βII-spectrin, that defines a boundary limiting ankG to the proximal axon. Experimentally moving this boundary altered the length of ankG staining in the proximal axon, whereas disruption of the boundary through silencing of ankB, αII-spectrin, or βII-spectrin expression blocked AIS assembly and permitted ankG to redistribute throughout the distal axon. In support of an essential role for the distal cytoskeleton in ankG clustering, we also found that αII and βII-spectrin-deficient mice had disrupted AIS. Thus, the distal axonal cytoskeleton functions as an intra-axonal boundary restricting ankG to the AIS.

We investigated CD45RA and CCR7 expression in CD4+ and CD8+ subsets of cerebrospinal fluid (CSF) lymphocytes, both immediately ex vivo and after stimulation, from 134 patients with a variety of inflammatory and non-inflammatory neurological diseases. Most inflammatory diseases had a higher CD4+:CD8+ ratio and higher percentage of effector memory T cells (T(EM)) than non-inflammatory controls, excluding active infection. Moreover, we found that patients with highly elevated cell counts in the CSF tended to have a lower percentage of central memory T cells (T(CM)) than patients with low or absent pleocytosis, with a concomitant increase in T(EM). We also found that samples with elevated IgG index or presence of oligoclonal bands had a significantly higher CD4+:CD8+ ratio than normal samples, consistent with increased CD4+ help for intrathecal IgG synthesis by B cells.

Primary resistance to pathogens is reliant on both basal and inducible immune defenses. To date, research has focused upon inducible innate immune responses. In contrast to resistance via cytokine induction, basal defense mechanisms are less evident. Here we showed that the antiviral protein kinase R (PKR) inhibited the key actin-modifying protein gelsolin to regulate actin dynamics and control cytoskeletal cellular functions under homeostatic conditions. Through this mechanism, PKR controlled fundamental innate immune, actin-dependent processes that included membrane ruffling and particle engulfment. Accordingly, PKR counteracted viral entry into the cell. These findings identify a layer of host resistance, showing that the regulation of actin-modifying proteins during the innate immune response bolsters first-line defense against intracellular pathogens and has a sustained effect on virus production. Moreover, these data provide proof of principle for a concept in which the cell cytoskeleton could be targeted to elicit broad antiviral protection.

Cell-type-specific control of gene expression is critical for the development of multicellular organisms. To investigate the mechanisms which underlie this, we have studied the regulation of the model genes Mdc and Il12b, whose stimulus-induced expression is tightly restricted to specific cells of the immune system. Surprisingly, we find that neither the promoter nor the enhancer sequences of these genes are sufficient to direct this cell-type specificity. Instead, the activities of upstream enhancers are repressed in nonexpressing cells by high levels of trimethylated H3K9 in their flanking regions. Genome-wide analysis indicates that this manner of regulation is shared by numerous enhancers of cell-type-specific genes. In dendritic cells and macrophages, the stimulus-induced demethylase Jmjd2d controls H3K9me3 levels at these regions, and is thereby required for Mdc and Il12b transcription. By experimentally assaying multiple enhancers in a variety of cell types, we show that regulation by H3K9me3 is a widely used mechanism which imparts specificity to the activities of otherwise broadly functional enhancers.

The role of speech production mechanisms in difficult speech comprehension is the subject of on-going debate in speech science. Two Activation Likelihood Estimation (ALE) analyses were conducted on neuroimaging studies investigating difficult speech comprehension or speech production. Meta-analysis 1 included 10 studies contrasting comprehension of less intelligible/distorted speech with more intelligible speech. Meta-analysis 2 (21 studies) identified areas associated with speech production. The results indicate that difficult comprehension involves increased reliance of cortical regions in which comprehension and production overlapped (bilateral anterior Superior Temporal Sulcus (STS) and anterior Supplementary Motor Area (pre-SMA)) and in an area associated with intelligibility processing (left posterior MTG), and second involves increased reliance on cortical areas associated with general executive processes (bilateral anterior insulae). Comprehension of distorted speech may be supported by a hybrid neural mechanism combining increased involvement of areas associated with general executive processing and areas shared between comprehension and production.

Deiters cells extend from the basilar membrane to the reticular lamina and, together with pillar cells and outer hair cells, structurally define the micro-architecture of the organ of Corti. Studying vibrotome sections of the mouse organ of Corti with confocal and scanning electron microscopy we found that the basal pole of every Deiters cell, independently of their position in the organ of Corti and along the cochlear spiral, attached to the basilar membrane within a 15.1 ± 0.3 μm-wide stripe running the length of the cochlear spiral adjacent to the row of outer pillar cells. All Deiters cells' basal poles had similar diameter and general morphology, and distributed on the stripe in a precise arrangement with a center-to-center distance of 7.1 ± 0.3 μm between neighbor cells of the same row and 5.9 ± 0.4 μm for neighbor cells in adjacent rows. Complete detachment of Deiters cells revealed an elliptical imprint on the top surface of the basilar membrane consisting of a smaller central structure with a very smooth surface surrounded by a rougher area, suggesting the presence of two different anchoring junctions. These previously unidentified morphological features of Deiters cells could be critical for the mechanical response of the organ of Corti.

Inflammation is associated with DNA damage, cellular senescence, and aging. Cessation of the inflammatory cytokine response is mediated in part through cytokine mRNA degradation facilitated by RNA-binding proteins, including AUF1. We report a major function of AUF1-it activates telomerase expression, suppresses cellular senescence, and maintains normal aging. AUF1-deficient mice undergo striking telomere erosion, markedly increased DNA damage responses at telomere ends, pronounced cellular senescence, and rapid premature aging that increases with successive generations, which can be rescued in AUF1 knockout mice and their cultured cells by resupplying AUF1 expression. AUF1 binds and strongly activates the transcription promoter for telomerase catalytic subunit Tert. In addition to directing inflammatory cytokine mRNA decay, AUF1 destabilizes cell-cycle checkpoint mRNAs, preventing cellular senescence. Thus, a single gene, AUF1, links maintenance of telomere length and normal aging to attenuation of inflammatory cytokine expression and inhibition of cellular senescence.

Recently, specific oxidized linoleic acid metabolites (OLAMs) have been identified as transient receptor potential vanilloid 1 (TRPV1) channel agonists that contribute to inflammatory and heat hyperalgesia mechanisms, yet the specific mechanism responsible for OLAM synthesis in sensory neurons is unknown. Here, we use molecular, anatomical, calcium imaging, and perforated patch electrophysiology methods to demonstrate the specific involvement of cytochrome P450 enzymes (CYPs) in the oxidation of linoleic acid leading to neuronal activation and show that this is enhanced under inflammatory conditions. Additional studies evaluated CYP expressions in the native rat trigeminal ganglia (TG) tissue and cultures as well as changes in their expression pattern following the induction of peripheral inflammation. Fourteen of 20 candidate transcripts were detected in native TG, and 7 of these displayed altered expression under cultured conditions. Moreover, complete Freund's adjuvant-induced inflammation of vibrissal pad selectively increased expression of CYP3A23/3A1 and CYP2J4 transcripts in TG. In situ hybridization studies demonstrated broad expression pattern of CYP3A23/3A1 and CYP2J4 within TG neurons. Anatomical studies characterized the expression of CYP3A1 and the CYP2J families within TG sensory neurons, including those with TRPV1, with about half of all TRPV1-positive neurons showing more prominent CYP3A1 and CYP2J expression. Together, these findings show that CYP enzymes play a primary role in mediating linoleic acid-evoked activation of sensory neurons and furthermore, implicate the involvement of specific CYPs as contributing to the formation of OLAMs that act as TRPV1 agonists within this subpopulation of nociceptors.

Lactobacillus plantarum is a facultative heterofermentative lactic acid bacterium widely used in the production of most fermented food due to its ability to thrive in several environmental niches, including the human gut. In order to cope with different growth conditions, it has developed complex molecular response mechanisms, characterized by the induction of a large set of proteins mainly regulated by HrcA and CtsR repressors as well as by global regulators such as carbon catabolite control protein A (CcpA). In this study, the role of CcpA in the regulation of growth under anaerobiosis and aerobiosis, and the adaptation to aeration in L. plantarum WCFS1 were comprehensively investigated by differential proteomics. The inactivation of ccpA, in both growth conditions, significantly changed the expression level of 76 proteins, mainly associated with carbohydrate and energy metabolism, membrane transport, nucleotide metabolism, protein biosynthesis and folding. The role of CcpA as pleiotropic regulator was particularly evident at the shift from homolactic fermentation to mixed fermentation. Proteomic results also indicated that the mutant strain was more responsive to aerobic growth condition.