Preferential binding to elk-1 by sle-associated il10 risk allele upregulates il10 expression

Immunoregulatory cytokine interleukin-10 (IL-10) is elevated in sera from patients with systemic lupus erythematosus (SLE) correlating with disease activity. The established association of IL10 with SLE and other autoimmune diseases led us to fine map causal variant(s) and to explore underlying mechanisms. We assessed 19 tag SNPs, covering the IL10 gene cluster including IL19, IL20 and IL24, for association with SLE in 15,533 case and control subjects from four ancestries. The previously reported IL10 variant, rs3024505 located at 1 kb downstream of IL10, exhibited the strongest association signal and was confirmed for association with SLE in European American (EA) (P = 2.7×10−8, OR = 1.30), but not in non-EA ancestries. SNP imputation conducted in EA dataset identified three additional SLE-associated SNPs tagged by rs3024505 (rs3122605, rs3024493 and rs3024495 located at 9.2 kb upstream, intron 3 and 4 of IL10, respectively), and SLE-risk alleles of these SNPs were dose-dependently associated with elevated levels of IL10 mRNA in PBMCs and circulating IL-10 protein in SLE patients and controls. Using nuclear extracts of peripheral blood cells from SLE patients for electrophoretic mobility shift assays, we identified specific binding of transcription factor Elk-1 to oligodeoxynucleotides containing the risk (G) allele of rs3122605, suggesting rs3122605 as the most likely causal variant regulating IL10 expression. Elk-1 is known to be activated by phosphorylation and nuclear localization to induce transcription. Of interest, phosphorylated Elk-1 (p-Elk-1) detected only in nuclear extracts of SLE PBMCs appeared to increase with disease activity. Co-expression levels of p-Elk-1 and IL-10 were elevated in SLE T, B cells and monocytes, associated with increased disease activity in SLE B cells, and were best downregulated by ERK inhibitor. Taken together, our data suggest that preferential binding of activated Elk-1 to the IL10 rs3122605-G allele upregulates IL10 expression and confers increased risk for SLE in European Americans.

Genesis and expansion of metazoan transcription factor gene classes

We know little about the genomic events that led to the advent of a multicellular grade of organization in animals, one of the most dramatic transitions in evolution. Metazoan multicellularity is correlated with the evolution of embryogenesis, which presumably was underpinned by a gene regulatory network reliant on the differential activation of signaling pathways and transcription factors. Many transcription factor genes that play critical roles in bilaterian development largely appear to have evolved before the divergence of cnidarian and bilaterian lineages. In contrast, sponges seem to have a more limited suite of transcription factors, suggesting that the developmental regulatory gene repertoire changed markedly during early metazoan evolution. Using whole- genome information from the sponge Amphimedon queenslandica, a range of eumetazoans, and the choanoflagellate Monosiga brevicollis, we investigate the genesis and expansion of homeobox, Sox, T- box, and Fox transcription factor genes. Comparative analyses reveal that novel transcription factor domains ( such as Paired, POU, and T- box) arose very early in metazoan evolution, prior to the separation of extant metazoan phyla but after the divergence of choanoflagellate and metazoan lineages. Phylogenetic analyses indicate that transcription factor classes then gradually expanded at the base of Metazoa before the bilaterian radiation, with each class following a different evolutionary trajectory. Based on the limited number of transcription factors in the Amphimedon genome, we infer that the genome of the metazoan last common ancestor included fewer gene members in each class than are present in extant eumetazoans. Transcription factor orthologues present in sponge, cnidarian, and bilaterian genomes may represent part of the core metazoan regulatory network underlying the origin of animal development and multicellularity.

Comparative expression analysis of transcription factor genes in the endostyle of invertebrate chordates

The endostyle of invertebrate chordates is a pharyngeal organ that is thought to be homologous with the follicular thyroid of vertebrates. Although thyroid-like features such as iodine-concentrating and peroxidase activities are located in the dorsolateral part of both ascidian and amphioxus endostyles, the structural organization and numbers of functional units are different. To estimate phylogenetic relationships of each functional zone with special reference to the evolution of the thyroid, we have investigated, in ascidian and amphioxus, the expression patterns of thyroid-related transcription factors such as TTF-2/MoxE4 and Pax2/5/8, as well as the forkhead transcription factors FoxQ1 and FoxA. Comparative gene expression analyses depicted an overall similarity between ascidians and amphioxus endostyles, while differences in expression patterns of these genes might be specifically related to the addition or elimination of a pair of glandular zones. Expressions of Ci-FoxE and BbFoxE4 suggest that the ancestral FoxE class might have been recruited for the formation of thyroid-like region in a possible common ancestor of chordates. Furthermore, coexpression of FoxE4, Pax2/5/8, and TPO in the dorsolateral part of both ascidian and amphioxus endostyles suggests that genetic basis of the thyroid function was already in place before the vertebrate lineage. (c) 2005 Wiley-Liss, Inc.

RE1 silencing transcription factor/neuron-restrictive silencing factor regulates expansion of adult mouse subventricular zone-derived neural stem/progenitor cells in vitro

Adult neural stem cell (aNSC) activity is tuned by external stimuli through the recruitment of transcription factors. This study examines the RE1 silencing transcription factor (REST) in neural stem/progenitor cells isolated from the subventricular zone of adult mouse brain and provides the first extensive characterization of REST-mediated control of the cellular and molecular properties. This study shows that REST knockdown affects the capacity of progenitor cells to generate neurospheres, reduces cell proliferation, and triggers cell differentiation despite the presence of growth factors. Genome- and transcriptome-wide analyses show that REST binding sites are significantly enriched in genes associated with synaptic transmission and nervous system development and function. Seeking candidate regulators of aNSC function, this study identifies a member of the bone morphogenetic protein (BMP) family, BMP6, the mRNA and protein of which increased after REST knockdown. The results of this study extend previous findings, demonstrating a reciprocal control of REST expression by BMPs. Administration of exogenous BMP6 inhibits aNSC proliferation and induces the expression of the astrocytic marker glial fibrillary acidic protein, highlighting its antimitogenic and prodifferentiative effects. This study suggests that BMP6 produced in a REST-regulated manner together with other signals can contribute to regulation of NSC maintenance and fate. © 2015 Wiley Periodicals, Inc.

Association of soluble tumor necrosis factor receptors 1 and 2 with nephropathy, cardiovascular events, and total mortality in type 2 diabetes

AIMS/HYPOTHESIS: Soluble tumor necrosis factor receptors 1 and 2 (sTNFR1 and sTNFR2) contribute to experimental diabetic kidney disease, a condition with substantially increased cardiovascular risk when present in patients. Therefore, we aimed to explore the levels of sTNFRs, and their association with prevalent kidney disease, incident cardiovascular disease, and risk of mortality independently of baseline kidney function and microalbuminuria in a cohort of patients with type 2 diabetes. In pre-defined secondary analyses we also investigated whether the sTNFRs predict adverse outcome in the absence of diabetic kidney disease. METHODS: The CARDIPP study, a cohort study of 607 diabetes patients [mean age 61 years, 44 % women, 45 cardiovascular events (fatal/non-fatal myocardial infarction or stroke) and 44 deaths during follow-up (mean 7.6 years)] was used. RESULTS: Higher sTNFR1 and sTNFR2 were associated with higher odds of prevalent kidney disease [odd ratio (OR) per standard deviation (SD) increase 1.60, 95 % confidence interval (CI) 1.32-1.93, p < 0.001 and OR 1.54, 95 % CI 1.21-1.97, p = 0.001, respectively]. In Cox regression models adjusting for age, sex, glomerular filtration rate and urinary albumin/creatinine ratio, higher sTNFR1 and sTNFR2 predicted incident cardiovascular events [hazard ratio (HR) per SD increase, 1.66, 95 % CI 1.29-2.174, p < 0.001 and HR 1.47, 95 % CI 1.13-1.91, p = 0.004, respectively]. Results were similar in separate models with adjustments for inflammatory markers, HbA1c, or established cardiovascular risk factors, or when participants with diabetic kidney disease at baseline were excluded (p < 0.01 for all). Both sTNFRs were associated with mortality. CONCLUSIONS/INTERPRETATIONS: Higher circulating sTNFR1 and sTNFR2 are associated with diabetic kidney disease, and predicts incident cardiovascular disease and mortality independently of microalbuminuria and kidney function, even in those without kidney disease. Our findings support the clinical utility of sTNFRs as prognostic markers in type 2 diabetes.

PTEN Directly Activates the Actin Depolymerization Factor Cofilin-1 During PGE(2)-Mediated Inhibition of Phagocytosis of Fungi

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evaluation of transformation growth factor beta(1), interleukin-10, and interferon-gamma in male symptomatic and asymptomatic dogs naturally infected by Leishmania (Leishmania) chagasi

The aims of this study were to evaluate the immunomodulatory role of TGF-beta(1), 1L-10, and INF-gamma in spleen and liver extracts and supernatant cultures of white spleen cells from male symptomatic and asymptomatic dogs, naturally infected by Leishmania (Leishmania) chagasi. Thirty dogs from Aracatuba, São Paulo, Brazil, an endemic leishmaniosis area, were selected by positive ELISA serological reaction for Leishmania sp. and divided into two groups: asymptomatic (n=15) and symptomatic (n=15) consisting of animals with at least three characteristic signs (fever, dermatitis, lymphoadenopathy, onychogryphosis, weight loss, cachex a, locomotion problems, conjunctivitis, epistaxis, hepatosplenomegaly, edema, and apathy). After euthanasia, spleen and liver fragments were collected for ex vivo quantification of TGF-beta(1), IL-10, and INF-gamma. Naturally active in vitro produced TGF-beta(1) was also evaluated in spleen cell culture supernatant. Spleen and liver extract of asymptomatic dogs had higher mean TGF-beta(1) levels than symptomatic dogs. High concentrations of IL-10 were found in spleen, and mainly in liver extract of both groups. Higher INF-gamma concentrations were found in spleen extracts of symptomatic dogs, and in liver extracts of asymptomatic dogs. Extract of this cytokire was lower in spleen extract. Although INF-gamma is being produced in canine infection, mean levels of TGF-beta(1) and IL-10 from spleen and liver extracts were quantitatively much higher; suggesting that immune response in both asymptomatic and symptomatic dogs A as predominantly type Th2. (c) 2006 Elsevier B.V. All rights reserved.

Influence of the dopaminergic system, CREB, and transcription factor-B on cocaine neurotoxicity

Cocaine is a widely used drug and its abuse is associated with physical, psychiatric and social problems. Abnormalities in newborns have been demonstrated to be due to the toxic effects of cocaine during fetal development. The mechanism by which cocaine causes neurological damage is complex and involves interactions of the drug with several neurotransmitter systems, such as the increase of extracellular levels of dopamine and free radicals, and modulation of transcription factors. The aim of this review was to evaluate the importance of the dopaminergic system and the participation of inflammatory signaling in cocaine neurotoxicity. Our study showed that cocaine activates the transcription factors NF-κB and CREB, which regulate genes involved in cellular death. GBR 12909 (an inhibitor of dopamine reuptake), lidocaine (a local anesthetic), and dopamine did not activate NF-κB in the same way as cocaine. However, the attenuation of NF-κB activity after the pretreatment of the cells with SCH 23390, a D1 receptor antagonist, suggests that the activation of NF-κB by cocaine is, at least partially, due to activation of D1 receptors. NF-κB seems to have a protective role in these cells because its inhibition increased cellular death caused by cocaine. The increase in BDNF (brain-derived neurotrophic factor) mRNA can also be related to the protective role of both CREB and NF-κB transcription factors. An understanding of the mechanisms by which cocaine induces cell death in the brain will contribute to the development of new therapies for drug abusers, which can help to slow down the progress of degenerative processes.

Macrophage Dectin-1 Expression Is Controlled by Leukotriene B-4 via a GM-CSF/PU.1 Axis

Pattern recognition receptors for fungi include dectin-1 and mannose receptor, and these mediate phagocytosis, as well as production of cytokines, reactive oxygen species, and the lipid mediator leukotriene B-4 (LTB4). The influence of G protein-coupled receptor ligands such as LTB4 on fungal pattern recognition receptor expression is unknown. In this study, we investigated the role of LTB4 signaling in dectin-1 expression and responsiveness in macrophages. Genetic and pharmacologic approaches showed that LTB4 production and signaling through its high-affinity G protein-coupled receptor leukotriene B4 receptor 1 (BLT1) direct dectin-1-dependent binding, ingestion, and cytokine production both in vitro and in vivo. Impaired responses to fungal glucans correlated with lower dectin-1 expression in macrophages from leukotriene (LT)- and BLT1-deficent mice than their wildtype counterparts. LTB4 increased the expression of the transcription factor responsible for dectin-1 expression, PU.1, and PU.1 small interfering RNA abolished LTB4-enhanced dectin-1 expression. GM-CSF controls PU.1 expression, and this cytokine was decreased in LT-deficient macrophages. Addition of GM-CSF to LT-deficient cells restored expression of dectin-1 and PU.1, as well as dectin-1 responsiveness. In addition, LTB4 effects on dectin-1, PU.1, and cytokine production were blunted in GM-CSF-/- macrophages. Our results identify LTB4-BLT1 signaling as an unrecognized controller of dectin-1 transcription via GM-CSF and PU.1 that is required for fungi-protective host responses. The Journal of Immunology, 2012, 189: 906-915.

CLONING AND EXPRESSION ANALYSIS OF ALLOGRAFT INFLAMMATORY FACTOR TYPE 1 IN COELOMOCYTES OF ANTARCTIC SEA URCHIN (STERECHINUS NEUMAYERI)

We have cloned and characterized for the first time an allograft inflammatory factor 1 (Sn-AIF-1) from the Antarctic sea urchin. We report the cloning of Sn-AIF-1 cDNA and the characterization of its expression in coelomocytes after a bacterial challenge. The cDNA Sn-AIF-1 has a size of 608 bp and encodes a polypeptide of 151 aa. The deduced amino acid sequence has a putative size of 17.430 Da, an isoelectric point of 4.92, and shows 2 elongation factor handlike motifs that normally bind calcium ions. BLAST analysis revealed close matches with other known AIF-1. The deduced amino acid sequence of Sn-AIF-1 showed high homology with AIF-1 in vertebrates such as fish, mice, and humans; and in the case of invertebrates, the major degree of identity (55%) was with a predicted sequence of the purple sea urchin AIF-1, and 52% corresponded to a sponge. Expression of Sn-AIF-1 mRNA was analyzed by qPCR. Sn-AIF-1 mRNA expression was measured from coelomocytes after a bacterial challenge using RT-PCR and revealed that the gene was upregulated after 24 h. Sn-AIF-1 could participate in the inflammatory response, particularly in the activation of coelomocytes and their survival.

PTEN Directly Activates the Actin Depolymerization Factor Cofilin-1 During PGE(2)-Mediated Inhibition of Phagocytosis of Fungi

Macrophage ingestion of the yeast Candida albicans requires its recognition by multiple receptors and the activation of diverse signaling programs. Synthesis of the lipid mediator prostaglandin E-2 (PGE(2)) and generation of cyclic adenosine monophosphate (cAMP) also accompany this process. Here, we characterized the mechanisms underlying PGE(2)-mediated inhibition of phagocytosis and filamentous actin (F-actin) polymerization in response to ingestion of C. albicans by alveolar macrophages. PGE(2) suppressed phagocytosis and F-actin formation through the PGE(2) receptors EP2 and EP4, cAMP, and activation of types I and II protein kinase A. Dephosphorylation and activation of the actin depolymerizing factor cofilin-1 were necessary for these inhibitory effects of PGE(2). PGE(2)-dependent activation of cofilin-1 was mediated by the protein phosphatase activity of PTEN (phosphatase and tensin homolog deleted on chromosome 10), with which it directly associated. Because enhanced production of PGE(2) accompanies many immunosuppressed states, the PTEN-dependent pathway described here may contribute to impaired antifungal defenses.

Molecular Characterization of the Putative Transcription Factor SebA Involved in Virulence in Aspergillus fumigatus

Aspergillus fumigatus is a major opportunistic pathogen and allergen of mammals. Nutrient sensing and acquisition mechanisms, as well as the capability to cope with different stressing conditions, are essential for A. fumigatus virulence and survival in the mammalian host. This study characterized the A. fumigatus SebA transcription factor, which is the putative homologue of the factor encoded by Trichoderma atroviride seb1. The Delta sebA mutant demonstrated reduced growth in the presence of paraquat, hydrogen peroxide, CaCl2, and poor nutritional conditions, while viability associated with sebA was also affected by heat shock exposure. Accordingly, SebA:GFP (SebA:green fluorescent protein) was shown to accumulate in the nucleus upon exposure to oxidative stress and heat shock conditions. In addition, genes involved in either the oxidative stress or heat shock response had reduced transcription in the Delta sebA mutant. The A. fumigatus Delta sebA strain was attenuated in virulence in a murine model of invasive pulmonary aspergillosis. Furthermore, killing of the Delta sebA mutant by murine alveolar macrophages was increased compared to killing of the wild-type strain. A. fumigatus SebA plays a complex role, contributing to several stress tolerance pathways and growth under poor nutritional conditions, and seems to be integrated into different stress responses.