Xiaoting Chen

Multi-species, multi-transcription factor binding highlights conserved control of tissue-specific biological pathways

As exome sequencing gives way to genome sequencing, the need to interpret the function of regulatory DNA becomes increasingly important. To test whether evolutionary conservation of cis-regulatory modules (CRMs) gives insight into human gene regulation, we determined transcription factor (TF) binding locations of four liver-essential TFs in liver tissue from human, macaque, mouse, rat, and dog. Approximately, two thirds of the TF-bound regions fell into CRMs. Less than half of the human CRMs were found as a CRM in the orthologous region of a second species. Shared CRMs were associated with liver pathways and disease loci identified by genome-wide association studies. Recurrent rare human disease causing mutations at the promoters of several blood coagulation and lipid metabolism genes were also identified within CRMs shared in multiple species. This suggests that multi-species analyses of experimentally determined combinatorial TF binding will help identify genomic regions critical for tissue-specific gene control. DOI: http://dx.doi.org/10.7554/eLife.02626.001

Interleukin-17 enhances immunosuppression by mesenchymal stem cells

IL-17 is one of the most potent and most actively investigated proinflammatory cytokines. In this study, we examined the effect of IL-17 on mesenchymal stem cells (MSCs) under the influence of inflammatory cytokines. Ironically, IL-17 dramatically enhanced the immunosuppressive effect of MSCs induced by IFNγ and TNFα, revealing a novel role of IL-17 in immunosuppression. Interestingly, we found that this action of IL-17 was dependent on the promoted expression of a key immune suppressive molecule, inducible nitric oxide synthase (iNOS), in MSCs. In a concanavalin A (ConA)-induced hepatitis mouse model, we found that IL-17 also enhanced the in vivo immunosuppressive effect of MSCs in an iNOS-dependent manner. Moreover, this promoting effect of IL-17 was found to be exerted through enhancing mRNA stability by modulating the protein level of ARE/poly(U)-binding/degradation factor 1 (AUF1), a well-known factor that promotes mRNA decay. In auf1−/− MSCs, IFNγ and TNFα could induce maximal immunosuppressive effect, both in vitro and in vivo, without the need for IL-17. Thus, our studies demonstrated that in the presence of MSCs, IL-17 promotes immunosuppression.

Topoisomerase 1 inhibition suppresses inflammatory genes and protects from death by inflammation

Unwinding DNA and unleasing inflammation Fighting infections often comes with collateral damage, which sometimes can be deadly. For instance, in septic shock, the overwhelming release of inflammatory mediators drives multi-organ failure. Rialdi et al. now report a potential new therapeutic target for controlling excessive inflammation: the DNA unwinding enzyme topoisomerase I (Top1) (see the Perspective by Pope and Medzhitov). Upon infection, Top1 specifically localizes to the promoters of pathogen-induced genes and promotes their transcription by helping to recruit RNA polymerase II. Pharmacological inhibition of Top1 in a therapeutic setting increased survival in several mouse models of severe microbially induced inflammation. Science, this issue p. 10.1126/science.aad7993; see also p. 1058 Depletion or chemical inhibition of Top1 suppresses the host response against influenza and Ebola viruses, as well as bacterial products. INTRODUCTION Infection causes inflammation, which contributes to pathogen clearance and organismal survival. The balance between the intensity and resolution of an inflammatory response is key for the fitness of the organism. Sepsis, for example, is a life-threatening condition caused by an excessive host response to infection, which in turn leads to multi-organ failure and death. Worldwide, millions of people each year succumb to sepsis. With an overall mortality rate of 20 to 50%, sepsis is the 10th leading cause of death (more than HIV and breast cancer) in the United States, according to the Centers for Disease Control and Prevention. Estimates indicate that 250,000 to 500,000 people die from sepsis annually in the United States. Children and the elderly are especially vulnerable to sepsis; it is the most common cause of death in infants and children. Childhood pneumonia, often caused by virus-bacteria co-infection, leads to septic shock and lung destruction. This occurs after bacterial invasion even in the presence of an appropriate antibiotic therapy. Finding remedies to treat sepsis and diseases associated with detrimental acute inflammatory reactions is thus pivotal for humankind. RATIONALE We reasoned that if excessive inflammation in response to infection leads to lethal consequences, dampening inflammation could be advantageous for the host. At least two strategies could be used to suppress inflammatory responses associated with infection. One is indirect and targets the pathogen (antibiotics). The second one, which we used, directly acts on the host response itself. In such a strategy, the suppression of acute inflammation would bypass the fatal outcome associated with overt inflammation and would “buy time” to allow the host immune response to eliminate the pathogen. After microbial invasion, many steps could be targeted between the early phases of the cellular response (sensing of the pathogen and signal transduction) and the information flow from DNA to RNA to proteins that act as inflammatory mediators (i.e., cytokines). We decided to identify and chemically inhibit cellular factors that act at the DNA (chromatin) level and play a primary role in activating the expression of inflammatory genes. RESULTS We found that chemical inhibition of topoisomerase 1 (Top1), an enzyme that unwinds DNA, suppresses the expression of infection-induced genes with little to no effect on housekeeping gene expression and without cellular damage. In vitro, depletion or chemical inhibition of Top1 in epithelial cells and macrophages suppresses the host response against influenza and Ebola viruses as well as bacterial products. At the mechanistic level, as shown by chemical genetics and epigenetic approaches, Top1 inhibition primarily suppresses RNA polymerase II (RNAPII) activity at pathogen-associated molecular pattern (PAMP)–induced genes. These genes require SWI/SNF chromatin remodeling for activation and display unique genetic and epigenetic features, such as the presence of IRF3 binding sites, low basal levels of RNAPII, histone H3 Lys27 acetylation marks, DNA hypersensitivity, and CpG islands. This gene “signature” of specificity was also validated using public data sets. In vivo, Top1 inhibition therapy rescued 70 to 90% mortality caused by exacerbated inflammation in three mouse models: acute bacteria infection, liver failure, and virus-bacteria co-infection. Strikingly, one to three doses of inhibitors were sufficient for the protective effect in all models, without overt side effects. CONCLUSION The inflammatory immune response against microbes is essential in protecting us against infections. In some cases, such as highly pathogenic and pandemic infections, the organism turns against itself and responds too acutely, with an excessive inflammation that can have fatal consequences. Our results suggest that a therapy based on Top1 inhibition could save millions of people affected by sepsis, pandemics, and many congenital deficiencies associated with acute inflammatory episodes and “cytokine storms.” CREDIT: RYGER/SHUTTERSTOCK The host innate immune response is the first line of defense against pathogens and is orchestrated by the concerted expression of genes induced by microbial stimuli. Deregulated expression of these genes is linked to the initiation and progression of diseases associated with exacerbated inflammation. We identified topoisomerase 1 (Top1) as a positive regulator of RNA polymerase II transcriptional activity at pathogen-induced genes. Depletion or chemical inhibition of Top1 suppresses the host response against influenza and Ebola viruses as well as bacterial products. Therapeutic pharmacological inhibition of Top1 protected mice from death in experimental models of lethal inflammation. Our results indicate that Top1 inhibition could be used as therapy against life-threatening infections characterized by an acutely exacerbated immune response.

The Human Transcription Factors

Transcription factors (TFs) recognize specific DNA sequences to control chromatin and transcription, forming a complex system that guides expression of the genome. Despite keen interest in understanding how TFs control gene expression, it remains challenging to determine how the precise genomic binding sites of TFs are specified and how TF binding ultimately relates to regulation of transcription. This review considers how TFs are identified and functionally characterized, principally through the lens of a catalog of over 1,600 likely human TFs and binding motifs for two-thirds of them. Major classes of human TFs differ markedly in their evolutionary trajectories and expression patterns, underscoring distinct functions. TFs likewise underlie many different aspects of human physiology, disease, and variation, highlighting the importance of continued effort to understand TF-mediated gene regulation.

MiR-125a targets effector programs to stabilize Treg-mediated immune homeostasis

Although different autoimmune diseases show discrete clinical features, there are common molecular pathways intimately involved. Here we show that miR-125a is downregulated in peripheral CD4(+) T cells of human autoimmune diseases including systemic lupus erythematosus and Crohns disease, and relevant autoimmune mouse models. miR-125a stabilizes both the commitment and immunoregulatory capacity of Treg cells. In miR-125a-deficient mice, the balance appears to shift from immune suppression to inflammation, and results in more severe pathogenesis of colitis and experimental autoimmune encephalomyelitis (EAE). The genome-wide target analysis reveals that miR-125a suppresses several effector T-cell factors including Stat3, Ifng and Il13. Using a chemically synthesized miR-125a analogue, we show potential to re-programme the immune homeostasis in EAE models. These findings point to miR-125a as a critical factor that controls autoimmune diseases by stabilizing Treg-mediated immune homeostasis.

Genetic Associations with Gestational Duration and Spontaneous Preterm Birth

BACKGROUND Despite evidence that genetic factors contribute to the duration of gestation and the risk of preterm birth, robust associations with genetic variants have not been identified. We used large data sets that included the gestational duration to determine possible genetic associations. METHODS We performed a genomewide association study in a discovery set of samples obtained from 43,568 women of European ancestry using gestational duration as a continuous trait and term or preterm (<37 weeks) birth as a dichotomous outcome. We used samples from three Nordic data sets (involving a total of 8643 women) to test for replication of genomic loci that had significant genomewide association (P<5.0×10‐8) or an association with suggestive significance (P<1.0×10‐6) in the discovery set. RESULTS In the discovery and replication data sets, four loci (EBF1, EEFSEC, AGTR2, and WNT4) were significantly associated with gestational duration. Functional analysis showed that an implicated variant in WNT4 alters the binding of the estrogen receptor. The association between variants in ADCY5 and RAP2C and gestational duration had suggestive significance in the discovery set and significant evidence of association in the replication sets; these variants also showed genomewide significance in a joint analysis. Common variants in EBF1, EEFSEC, and AGTR2 showed association with preterm birth with genomewide significance. An analysis of mother–infant dyads suggested that these variants act at the level of the maternal genome. CONCLUSIONS In this genomewide association study, we found that variants at the EBF1, EEFSEC, AGTR2, WNT4, ADCY5, and RAP2C loci were associated with gestational duration and variants at the EBF1, EEFSEC, and AGTR2 loci with preterm birth. Previously established roles of these genes in uterine development, maternal nutrition, and vascular control support their mechanistic involvement. (Funded by the March of Dimes and others.)

The interaction between mesenchymal stem cells and steroids during inflammation

Mesenchymal stem cells (MSCs) are believed to exert their regenerative effects through differentiation and modulation of inflammatory responses. However, the relationship between the severity of inflammation and stem cell-mediated tissue repair has not been formally investigated. In this study, we applied different concentrations of dexamethasone (Dex) to anti-CD3-activated splenocyte cultured with or without MSCs. As expected, Dex exhibited a classical dose-dependent inhibition of T-cell proliferation. Surprisingly, although MSCs also blocked T-cell proliferation, the presence of Dex unexpectedly showed a dose-dependent reversion of T-cell proliferation. This effect of Dex was found to be exerted through interfering STAT1 phosphorylation-prompted expression of inducible nitric oxide synthase (iNOS). Interestingly, inflammation-induced chemokines in MSCs was unaffected. To test the role of inflammation severity in stem cell-mediated tissue repair, we employed mice with carbon tetrachloride-induced advanced liver fibrosis and found that although MSCs alone were effective, concurrent administration of Dex abrogated the therapeutic effects of MSCs on fibrin deposition, serum levels of bilirubin, albumin, and aminotransferases, as well as T-lymphocyte infiltration, especially IFN-γ+CD4+ and IL-17A+CD4+T cells. Likewise, iNOS−/− MSCs, which produce chemokines but not nitric oxide under inflammatory conditions, are ineffective in treating advanced liver fibrosis. Therefore, inflammation has a critical role in MSC-mediated tissue repair. In addition, concomitant application of MSCs with steroids should be avoided.

DNA methylation dynamics during ex vivo differentiation and maturation of human dendritic cells

BackgroundDendritic cells (DCs) are important mediators of innate and adaptive immune responses, but the gene networks governing their lineage differentiation and maturation are poorly understood. To gain insight into the mechanisms that promote human DC differentiation and contribute to the acquisition of their functional phenotypes, we performed genome-wide base-resolution mapping of 5-methylcytosine in purified monocytes and in monocyte-derived immature and mature DCs.ResultsDC development and maturation were associated with a great loss of DNA methylation across many regions, most of which occurs at predicted enhancers and binding sites for known transcription factors affiliated with DC lineage specification and response to immune stimuli. In addition, we discovered novel genes that may contribute to DC differentiation and maturation. Interestingly, many genes close to demethylated CG sites were upregulated in expression. We observed dynamic changes in the expression of TET2, DNMT1, DNMT3A and DNMT3B coupled with temporal locus-specific demethylation, providing possible mechanisms accounting for the dramatic loss in DNA methylation.ConclusionsOur study is the first to map DNA methylation changes during human DC differentiation and maturation in purified cell populations and will greatly enhance the understanding of DC development and maturation and aid in the development of more efficacious DC-based therapeutic strategies.

Myeloid malignancies with chromosome 5q deletions acquire a dependency on an intrachromosomal NF-κB gene network.

Chromosome 5q deletions (del[5q]) are common in high-risk (HR) myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML); however, the gene regulatory networks that sustain these aggressive diseases are unknown. Reduced miR-146a expression in del(5q) HR MDS/AML and miR-146a(-/-) hematopoietic stem/progenitor cells (HSPCs) results in TRAF6/NF-κB activation. Increased survival and proliferation of HSPCs from miR-146a(low) HR MDS/AML is sustained by a neighboring haploid gene, SQSTM1 (p62), expressed from the intact 5q allele. Overexpression of p62 from the intact allele occurs through NF-κB-dependent feedforward signaling mediated by miR-146a deficiency. p62 is necessary for TRAF6-mediated NF-κB signaling, as disrupting the p62-TRAF6 signaling complex results in cell-cycle arrest and apoptosis of MDS/AML cells. Thus, del(5q) HR MDS/AML employs an intrachromosomal gene network involving loss of miR-146a and haploid overexpression of p62 via NF-κB to sustain TRAF6/NF-κB signaling for cell survival and proliferation. Interfering with the p62-TRAF6 signaling complex represents a therapeutic option in miR-146a-deficient and aggressive del(5q) MDS/AML.

Increased expression of the type I interferon-inducible gene, lymphocyte antigen 6 complex locus E, in peripheral blood cells is predictive of lupus activity in a large cohort of Chinese lupus patients:

Several studies by microarray analysis and real-time polymerase chain reaction (RT-PCR) reveal that type I interferon-inducible genes (IFIGs) are implicated in systemic lupus erythematosus (SLE). To find a potential clinical biomarker capable of monitoring lupus disease activity clinically, quantitative RT-PCR was used to identify transcript expression levels of 13 type I IFIGs in peripheral blood cells in 144 patients with SLE, 27 non-SLE patients and 60 healthy controls and then analyse connections between gene expression and disease activity. The expression levels of five type I IFIGs (LY6E, OAS3, IFIT4, OAS1 and OAS2) were significantly higher in the SLE group than in the healthy and non-SLE controls. LY6E gene that had highest expression was chosen to analyse the association of expression level with clinical features. Compared to low LY6E expression group, SLE patients with high LY6E expression had higher SLEDAI-2K score, increased 24 h urine protein and lower blood C3 complement. Active SLE patients had more elevated LY6E expression than stable patients. And LY6E expression levels in patients with SLE were strongly correlated with their SLEDAI-2K scores. Our results indicate that increased expression of LY6E gene in peripheral blood cells in patients with SLE is correlated with lupus activity and may be a useful, noninvasive biomarker for assessing SLE disease activity.