cAMP target sequences enhCRE and CNRE sense low-salt intake to increase human renin gene expression in vivo

This study aimed to assess the role of cAMP target sequences enhancer cAMP response element (enhCRE) and cAMP and overlapping negative response element (CNRE) in the control of human renin gene (REN) in vivo. enhCRE and CNRE were silenced by mutations in a 12.2-kb human renin promoter fused to LacZ reporter gene. This construct was used to generate transgenic mice (RENMut-LacZ). The expression of the transgene was correctly targeted to the juxtaglomerular portions of renal afferent arterioles which express endogenous mouse renin. Therefore, enhCRE and CNRE do not seem to be relevant for the control of the cell-specific expression of the human renin gene. The β-adrenoreceptor agonist isoproterenol (10?mg/kg/day, for 2?days) stimulated the endogenous renin, but not the LacZ mRNA expression. Treatment of RENMut-LacZ mice with the angiotensin converting enzyme inhibitor (enalapril 10?mg/kg/day, for 7?days) or their crossing to angiotensin receptor type 1a knockout mice led to increased renin and LacZ mRNA levels. Renin expression was upregulated by low-salt diet (0.03% NaCl, for 10?days) and downregulated by high-salt diet (4% NaCl, for 10?days). In contrast, low-salt diet did not influence, while high-salt diet inhibited the expression of LacZ. In summary, enhCRE and CNRE appear to be necessary for the transactivation of the human renin gene through β-adrenoreceptors and by low-salt diet. Our data also suggest that different intracellular mechanisms mediate the effect of low- and high-salt intake on renin expression in vivo.     

Implicit trustworthiness decisions: automatic coding of face properties in the human amygdala

Deciding whether an unfamiliar person is trustworthy is one of the most important decisions in social environments. We used functional magnetic resonance imaging to show that the amygdala is involved in implicit evaluations of trustworthiness of faces, consistent with prior findings. The amygdala response increased as perceived trustworthiness decreased in a task that did not demand person evaluation. More importantly, we tested whether this response is due to an individual's idiosyncratic perception or to face properties that are perceived as untrustworthy across individuals. The amygdala response was better predicted by consensus ratings of trustworthiness than by an individual's own judgments. Individual judgments accounted for little residual variance in the amygdala after controlling for the shared variance with consensus ratings. These findings suggest that the amygdala automatically categorizes faces according to face properties commonly perceived to signal untrustworthiness.     

Predicting political elections from rapid and unreflective face judgments

Here we show that rapid judgments of competence based solely on the facial appearance of candidates predicted the outcomes of gubernatorial elections, the most important elections in the United States next to the presidential elections. In all experiments, participants were presented with the faces of the winner and the runner-up and asked to decide who is more competent. To ensure that competence judgments were based solely on facial appearance and not on prior person knowledge, judgments for races in which the participant recognized any of the faces were excluded from all analyses. Predictions were as accurate after a 100-ms exposure to the faces of the winner and the runner-up as exposure after 250 ms and unlimited time exposure (Experiment 1). Asking participants to deliberate and make a good judgment dramatically increased the response times and reduced the predictive accuracy of judgments relative to both judgments made after 250 ms of exposure to the faces and judgments made within a response deadline of 2 s (Experiment 2). Finally, competence judgments collected before the elections in 2006 predicted 68.6% of the gubernatorial races and 72.4% of the Senate races (Experiment 3). These effects were independent of the incumbency status of the candidates. The findings suggest that rapid, unreflective judgments of competence from faces can affect voting decisions.     

Metals and health: a clinical toxicological perspective on tungsten and review of the literature

Tungsten and tungsten compounds are considered toxicologically relatively safe. Concern regarding the potential health and environmental effects of depleted uranium and lead in military applications has lead many countries to explore the possibility of applying toxicologically safer metals. Heavy metal tungsten alloy-based munitions have been therefore introduced as a replacement in munitions and as kinetic energy penetrators. Although the toxicological profiles of all these metals are well known, their internalization as embedded shrapnel may be considered a new route for long-term exposure. Studies in experimental animals and cell culture indicate that pellets based on heavy metal tungsten alloy possess carcinogenic potential previously unseen for depleted uranium and/or lead. Other metals in the tungsten alloy such as nickel or cobalt may contribute to such a risk. Accordingly, the long-term tungsten-related health risk is reason for concern. This article reviews toxicological and clinical literature and provides new perspectives on tungsten and tungsten-based alloys.     

Total ganglioside ablation at mouse motor nerve terminals alters neurotransmitter release level

Neuronal membrane gangliosides, forming a large family of sialylated glycosphingolipids, have been hypothesized to play important roles in synaptic transmission. We studied the ex vivo electrophysiological function of neuromuscular junctions of GM2/GD2‐synthase*GD3‐synthase compound null‐mutant mice after acute removal of GM3, the only remaining ganglioside in this mouse, by in vitro treatment with neuraminidase. We found 16% enhancement of the acetylcholine release per nerve impulse at low‐rate (0.3 Hz) nerve stimulation. Conversely, the treatment reduced the acetylcholine release evoked by high‐rate (40 Hz) nerve stimulation. Also, 25 ms paired‐pulse facilitation of endplate potentials was reduced by the neuraminidase‐treatment. These effects may indicate a modest modulatory influence of the negative electrical charges carried by the sialic acid molecules of gangliosides on the function of presynaptic Ca_v2.1 channels, affecting the magnitude and kinetics of the Ca²⁺ influx that induces neurotransmitter release from the motor nerve terminal. Our results show that gangliosides are to some extent involved in neurotransmission at the neuromuscular junction, but that their presence is not an absolute requirement in this process. Synapse 64:335–338, 2010. © 2009 Wiley‐Liss, Inc.     

A family study of complex chromosome rearrangement involving chromosomes 1, 8, and 11 and its reproductive consequences

Complex chromosome translocations are structural chromosomal rearrangements involving three or more chromosomes and more than two breakpoints. A complex chromosome rearrangement was detected in a phenotypically normal female patient that was referred to the hospital for genetic counseling due to reproductive failure. A cytogenetic evaluation was performed, according to standard method of chromosomal analysis, using G-banding technique. The patient’s karyotype showed a balanced complex chromosome rearrangement (BCCR) involving chromosomes 1, 8, and 11 with three breakpoints 1p31, 8q13, and 11q23. The karyotype designed according to ISCN (2013), is 46,XX,t(1;8;11)(p31;q13;q23) (8qter→8q13::1p31→1qter;8pter→8q13::11q23→11qter;11pter→11q23::1p31→1pter). Additionally, the proband’s mother and brother were tested, resulting in the same exact translocation. In this study, we describe all possible meiotic segregations regarding this translocation, as well as the clinical phenotypes which could arise, if unbalanced products of conception survive. This is a rare case of familial complex chromosome rearrangement, giving a view for its reproductive consequences.     

Germ-free housing conditions do not affect aortic root and aortic arch lesion size of late atherosclerotic low-density lipoprotein receptor-deficient mice

ABSTRACT

The microbiota has been linked to the development of atherosclerosis, but the functional impact of these resident bacteria on the lesion size and cellular composition of atherosclerotic plaques in the aorta has never been experimentally addressed with the germ-free low-density lipoprotein receptor-deficient (Ldlr^?/- ) mouse atherosclerosis model. Here, we report that 16 weeks of high-fat diet (HFD) feeding of hypercholesterolemic Ldlr^?/- mice at germ-free (GF) housing conditions did not impact relative aortic root plaque size, macrophage content, and necrotic core area. Likewise, we did not find changes in the relative aortic arch lesion size. However, late atherosclerotic GF Ldlr^?/- mice had altered inflammatory plasma protein markers and reduced smooth muscle cell content in their atherosclerotic root plaques relative to CONV-R Ldlr^?/- mice. Neither absolute nor relative aortic root or aortic arch plaque size correlated with age. Our analyses on GF Ldlr^?/- mice did not reveal a significant contribution of the microbiota in late aortic atherosclerosis.     

HDAC inhibitor reduces cytokine storm and facilitates induction of chimerism that reverses lupus in anti-CD3 conditioning regimen

In allogeneic hematopoietic cell transplantation (HCT), donor T cell-mediated graft versus host leukemia (GVL) and graft versus autoimmune (GVA) activity play critical roles in treatment of hematological malignancies and refractory autoimmune diseases. However, graft versus host disease (GVHD), which sometimes can be fatal, remains a major obstacle in classical HCT, where recipients are conditioned with total body irradiation or high-dose chemotherapy. We previously reported that anti-CD3 conditioning allows donor CD8(+) T cells to facilitate engraftment and mediate GVL without causing GVHD. However, the clinical application of this radiation-free and GVHD preventative conditioning regimen is hindered by the cytokine storm syndrome triggered by anti-CD3 and the high-dose donor bone marrow (BM) cells required for induction of chimerism. Histone deacetylase (HDAC) inhibitors such as suberoylanilide hydroxamic acid (SAHA) are known to induce apoptosis of cancer cells and reduce production of proinflammatory cytokines by nonmalignant cells. Here, we report that SAHA inhibits the proliferative and cytotoxic activity of anti-CD3-activated T cells. Administration of low-dose SAHA reduces cytokine production and ameliorates the cytokine storm syndrome triggered by anti-CD3. Conditioning with anti-CD3 and SAHA allows induction of chimerism with lower doses of donor BM cells in old nonautoimmune and autoimmune lupus mice. In addition, conditioning with anti-CD3 and SAHA allows donor CD8(+) T cell-mediated GVA activity to reverse lupus glomerulonephritis without causing GVHD. These results indicate that conditioning with anti-CD3 and HDAC inhibitors represent a radiation-free and GVHD-preventative regimen with clinical application potential.     

Comprehensive assessment of cancer missense mutation clustering in protein structures

Large-scale tumor sequencing projects enabled the identification of many new cancer gene candidates through computational approaches. Here, we describe a general method to detect cancer genes based on significant 3D clustering of mutations relative to the structure of the encoded protein products. The approach can also be used to search for proteins with an enrichment of mutations at binding interfaces with a protein, nucleic acid, or small molecule partner. We applied this approach to systematically analyze the PanCancer compendium of somatic mutations from 4,742 tumors relative to all known 3D structures of human proteins in the Protein Data Bank. We detected significant 3D clustering of missense mutations in several previously known oncoproteins including HRAS, EGFR, and PIK3CA. Although clustering of missense mutations is often regarded as a hallmark of oncoproteins, we observed that a number of tumor suppressors, including FBXW7, VHL, and STK11, also showed such clustering. Beside these known cases, we also identified significant 3D clustering of missense mutations in NUF2, which encodes a component of the kinetochore, that could affect chromosome segregation and lead to aneuploidy. Analysis of interaction interfaces revealed enrichment of mutations in the interfaces between FBXW7-CCNE1, HRAS-RASA1, CUL4B-CAND1, OGT-HCFC1, PPP2R1A-PPP2R5C/PPP2R2A, DICER1-Mg²⁺, MAX-DNA, SRSF2-RNA, and others. Together, our results indicate that systematic consideration of 3D structure can assist in the identification of cancer genes and in the understanding of the functional role of their mutations.To elucidate the genetic basis of cancer, efforts have been initiated to sequence the exomes or genomes of many human tumors. Among them are large-scale efforts such as The Cancer Genome Atlas (TCGA) (1) and the International Cancer Genome Consortium (ICGC) (2), as well as many smaller-scale projects. These efforts have collectively found millions of somatic mutations in virtually all human genes (the vast majority of which are nonfunctional or “passenger” mutations) across thousands of tumor samples (3). The amounts of available cancer sequencing data are growing rapidly and will continue to grow in the foreseeable future. We and others have developed computational methods to detect cancer-associated genes and functional mutations from such data, based on a significant overall burden of mutations or on significant positional clustering of mutations in the one-dimensional (1D) gene sequences, corresponding to mutational hotspots (3–6).For some cancer proteins, it has been observed that, although mutations may be distributed along the linear amino acid sequence, they tend to cluster in certain regions of the 3D structure, such as active sites. A clear example is KRAS, where particular missense mutations at the active site are positively selected in cancer because they disable the GTPase activity of the protein, locking it in its GTP-bound, active state, which promotes proliferation. As a result, recurrently mutated residues (e.g., G12, G13, I36, A59, Q61, K117, A146) tend to occur around the substrate-binding pocket of KRAS (Fig. 1). This and other individual examples of proteins showing 3D clustering of cancer missense mutations are sometimes used in the literature as supporting evidence for the involvement of those proteins in the disease or as a basis for functional hypotheses about the clustered mutations [e.g., EGFR (8), PIK3CA (9), DIS3 (10), SPOP (11), MRE11 (12), ERCC2 (13)]. Stehr et al. (14) and Ryslik et al. (15) assessed the structural clustering of missense mutations in 29 and 131 proteins, respectively, and demonstrated that taking into account 3D structural information can be helpful for identifying mutation hotspots in known cancer proteins or in new candidates.Open in a separate windowFig. 1.Spatial mutation clustering in KRAS. (A) Protein sequence of KRAS (Isoform 2B; UniProt: P01116-2) with mutated residues from the PanCancer data set (3) shown in red. Recurrent mutations (at least three samples) are shown in larger font and are annotated with position and number of samples with such mutations. Gray arcs between such residues are shown if their centroids are located closer than 13 Å between each other in the protein structure; arc width and label show the spatial distance in these cases (wider arcs corresponding to shorter distances). The C-terminal part of the protein sequence not covered by the structure in B is shown in smaller, gray font. (B) 3D structure of KRAS (gray) with substrate GDP (blue) bound to its active site (PDB ID code 4LUC) (7). Mutated residues are shown in red (recurrent mutations: sticks, nonrecurrent mutations: thin lines) and color intensity scales with the number of mutations per residue.Here, we seek to undertake comprehensive studies of 3D clustering of somatic missense mutations in cancer across all human proteins with available protein structures. Such integrative analysis may help to identify new cancer proteins that have been missed by other methods. In addition, it can help explain the functional roles of individual mutations based on their spatial location in the protein; for example, mutations that cluster at protein interaction interfaces may perturb key molecular interactions (16).