AHR is a tunable knob that controls HTLV-1 latency-reactivation switching.

Establishing latent infection but retaining the capability to reactivate in certain circumstance is an ingenious tactic for retroviruses to persist in vivo while evading host immune surveillance. Many evidences indicate that Human T-cell leukemia virus type 1 (HTLV-1) is not completely silent in vivo. However, signals that trigger HTLV-1 latency-reactivation switching remain poorly understood. Here, we show that aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor, plays a critical role in HTLV-1 plus-strand expression. Importantly, HTLV-1 reactivation could be tunably manipulated by modulating the level of AHR ligands. Mechanistically, activated AHR binds to HTLV-1 LTR dioxin response element (DRE) site (CACGCATAT) and drives plus-strand transcription. On the other hand, persistent activation of nuclear factor kappa B (NF-κB) pathway constitutes one key prerequisite for AHR overexpression in HTLV-1 infected T-cells, setting the stage for the advent of AHR signaling. Our findings suggest that HTLV-1 might achieve its reactivation in vivo when encountering environmental, dietary, microbial and metabolic cues that induce sufficient AHR signaling

Identification of a novel macrophage-related prognostic signature in colorectal cancer

Abstract Colorectal cancer (CRC) is one of the most prevalent and deadliest illnesses all around the world. Growing proofs demonstrate that tumor-associated macrophages (TAMs) are of critical importance in CRC pathogenesis, but their mechanisms remain yet unknown. The current research was designed to recognize underlying biomarkers associated with TAMs in CRC. We screened macrophage-related gene modules through WGCNA, selected hub genes utilizing the LASSO algorithm and COX regression, and established a model. External validation was performed by expression analysis using datasets GSE14333, GSE74602, and GSE87211. After validating the bioinformatics results using real-time quantitative reverse transcription PCR, we identified SPP1, C5AR1, MMP3, TIMP1, ADAM8 as potential biomarkers associated with macrophages in CRC

IL-8 Secreted from M2 Macrophages Promoted Prostate Tumorigenesis via STAT3/MALAT1 Pathway

Prostate cancer (PCa) is a major health problem in males. Metastasis-associated with lung adenocarcinoma transcript-1 (MALAT1), which is overexpressed in PCa tissue, is associated with physiological and pathological conditions of PCa. M2 macrophages are major immune cells abundant in the tumor microenvironment. However, it remains unknown whether M2 macrophages are involved in the effects or not, and molecular mechanisms of MALAT1 on PCa progression have not yet been comprehensively explored. Here we reported that, M2 macrophages (PMA/IL-4 treated THP1) induced MALAT1 expression in PCa cell lines. Knockdown MALAT1 expression level in PCa cell lines inhibited cellular proliferation, invasion, and tumor formation. Further mechanistic dissection revealed that M2 macrophages secreted IL-8 was sufficient to drive up MALAT1 expression level via activating STAT3 signaling pathway. Additional chromatin immunoprecipitation (ChIP) and luciferase reporter assays displayed that STAT3 could bind to the MALAT1 promoter region and transcriptionally stimulate the MALAT1 expression. In summary, our present study identified the IL-8/STAT3/MALAT1 axis as key regulators during prostate tumorigenesis and therefore demonstrated a new mechanism for the MALAT1 transcriptional regulation

A Crystal Structure of the Dengue Virus NS5 Protein Reveals a Novel Inter-domain Interface Essential for Protein Flexibility and Virus Replication

International audienceFlavivirus RNA replication occurs within a replication complex (RC) that assembles on ER membranes and comprises both non-structural (NS) viral proteins and host cofactors. As the largest protein component within the flavivirus RC, NS5 plays key enzymatic roles through its N-terminal methyltransferase (MTase) and C-terminal RNA-dependent-RNA polymerase (RdRp) domains, and constitutes a major target for antivirals. We determined a crystal structure of the full-length NS5 protein from Dengue virus serotype 3 (DENV3) at a resolution of 2.3 Å in the presence of bound SAH and GTP. Although the overall molecular shape of NS5 from DENV3 resembles that of NS5 from Japanese Encephalitis Virus (JEV), the relative orientation between the MTase and RdRp domains differs between the two structures, providing direct evidence for the existence of a set of discrete stable molecular conformations that may be required for its function. While the inter-domain region is mostly disordered in NS5 from JEV, the NS5 structure from DENV3 reveals a well-ordered linker region comprising a short 310 helix that may act as a swivel. Solution Hydrogen/Deuterium Exchange Mass Spectrometry (HDX-MS) analysis reveals an increased mobility of the thumb subdomain of RdRp in the context of the full length NS5 protein which correlates well with the analysis of the crystallographic temperature factors. Site-directed mutagenesis targeting the mostly polar interface between the MTase and RdRp domains identified several evolutionarily conserved residues that are important for viral replication, suggesting that inter-domain cross-talk in NS5 regulates virus replication. Collectively, a picture for the molecular origin of NS5 flexibility is emerging with profound implications for flavivirus replication and for the development of therapeutics targeting NS5

A schematic model for the divergent evolution of flaviviral NS5 proteins.

<p>The same color scheme as in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004682#ppat.1004682.g001" target="_blank">Fig. 1</a> is used: MTase is in yellow, RdRp fingers in green, palm in blue, thumb in salmon. The linker region 3₁₀ helix (residues 263–266) between the two domains is in orange. Active sites for MTase and RdRp are labelled with dotted tetragon and pentagon respectively. Linker residues and interface residues are labeled. A possible evolutionary pathway is presented: the MTase domain and RdRp domain originally existing as two separate proteins (left) became linked together to form the NS5 protein from an ancestral Flavivirus, possibly through gene fusion. This fusion promoted colocalization of both enzymatic activities and effectively increased the effective concentration of the proteins with respect to each other (middle panel). Following further (divergent) evolution, NS5 acquired different adaptive mutations and gave rise to the NS5 proteins now observed for various viruses, including DENV, JEV and possibly other flaviviruses) (right panel). Thus NS5 proteins from DENV and JEV may have different conformations and different allosteric mechanisms, in which the MTase and RdRp domain cross-talk to each other through unique interfaces specific to either DENV or JEV [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004682#ppat.1004682.ref072" target="_blank">72</a>].</p

Replication profiles of NS5 interface mutants.

<p><b>(A)</b> Renilla luciferase activities of DENV4 WT and mutant replicons. Equal amounts of replicon RNA (WT or mutants) were electroporated into BHK-21 cells. At the indicated time points, the transfected cells were lysed and assayed for luciferase activities. The y axis shows the log10 value of Renilla luciferase activity (RLU). Each data point is the average for three replicates, and error bars show the standard deviations. <b>(B)</b> 10μg <i>in vitro</i> transcribed infectious clone RNA was electroporated into BHK-21 cells and viral replication was monitored over a course of 5 days. Intracellular viral RNA replication as detected by qRT-PCR. The grey dotted line represents the background detection of uninfected cells. <b>(C)</b> Extracellular viral RNA in the supernatants detected by qRT-PCR. The grey dotted line denotes background signal of uninfected supernatant. <b>(D)</b> Plaque morphologies of WT and the mutants at 72 hours post electroporation. <b>(E)</b> IFA images showing dsRNA and NS5 co-staining and percentage infection of cells at 72 hours post electroporation.</p

Data collection and refinement statistics.

<p>*Statistics for the highest-resolution shell are shown in parentheses.</p><p>Data collection and refinement statistics.</p

Crystal structure of DENV3 NS5.

<p><b>(A)</b> Overall structure of the NS5 protein from DENV3 in cartoon representation viewing from the bottom of RdRp. MTase is in yellow, RdRp fingers in green, palm in blue, thumb in salmon. The linker helix (residues 263–267) between the two domains is in orange. GTP and co-factor SAH are shown as sticks and labelled. Zinc ions are shown as spheres. <b>(B)</b> View of the NS5 molecule from the top of RdRp, which is rotated by 180° around a vertical axis as in (A). Interface regions are boxed. <b>(C)</b> and <b>(D</b>) Close-up views of the interface between the MTase domain and RdRp domain as indicated in (B). Key residues for inter-domain interactions are shown as sticks and labeled. <b>(E)</b> Multiple sequence alignment of flavivirus NS5 proteins. Interface residues are highlighted in gray. The linker residues (263–272 in DENV3 NS5) are boxed. List of accession numbers for genes and proteins used for alignment: DENV3: gi|50347097|gb|AAT75224.1|; DENV1: gi|194338413|gb|ACF49259.1|; DENV2: gi|266813|sp|P29990.1|; DENV4: gi|425895219|gb|AFY10034.1|; JEV: gi|4416167|gb|AAD20233.1|; WNV: gi|607369775|gb|AHW48802.1|; YFV: gi|27735297|ref|NP_776009.1|; TBEV: gi|1709707|sp|Q01299.1|.</p