Analysis of MEFV exon methylation and expression patterns in familial Mediterranean fever

<p>Abstract</p> <p>Background</p> <p>MEFV mutations and decreased expression level of the gene are related to FMF pathology. DNA methylation at CpG islands is a well-known mechanism for transcriptional silencing. MEFV has a CpG island, spanning a part of the first intron and the whole of the second exon of the gene covering 998 bp region. Here, we tested the hypothesis that the MEFV transcript level in FMF patients correlates with its methylation level, and methylation, by allowing transcription silencing, has a role in FMF ethiopathogenesis.</p> <p>Methods</p> <p>The study group was composed of pediatric FMF patients (N = 51) and age-gender matched healthy controls (N = 21). The relative expression level of MEFV was assessed via quantitative real-time PCR (qRT-PCR) and bisulfite sequencing (BS) was performed to analyse the methylation level quantitatively.</p> <p>Results</p> <p>MEFV expression in FMF patients were decreased compared to healthy controls (<it>P </it>= 0.031). Methylation level of exon 2 of MEFV was found to be slightly higher in FMF patients compared to healthy controls (76% versus 74%) (<it>P </it>= 0.049). The expression level of the MEFV was negatively correlated with the methylation level of the CpG island in both FMF and healthy controls groups (cor = -0.29, <it>P </it>= 0.041) but more so in the FMF only group (cor = -0.36, <it>P </it>= 0.035).</p> <p>Conclusions</p> <p>In this study, the relation between reduced MEFV expression level and FMF was confirmed. Observed slight increase in methylation in FMF patients, and correlation of methylation with expression might be indicative of its role in FMF, however a larger dataset is needed to confirm our preliminary findings.</p

A comparative analysis of two conserved motifs in bacterial poly(A) polymerase and CCA-adding enzyme

Showing a high sequence similarity, the evolutionary closely related bacterial poly(A) polymerases (PAP) and CCA-adding enzymes catalyze quite different reactions—PAP adds poly(A) tails to RNA 3′-ends, while CCA-adding enzymes synthesize the sequence CCA at the 3′-terminus of tRNAs. Here, two highly conserved structural elements of the corresponding Escherichia coli enzymes were characterized. The first element is a set of amino acids that was identified in CCA-adding enzymes as a template region determining the enzymes' specificity for CTP and ATP. The same element is also present in PAP, where it confers ATP specificity. The second investigated region corresponds to a flexible loop in CCA-adding enzymes and is involved in the incorporation of the terminal A-residue. Although, PAP seems to carry a similar flexible region, the functional relevance of this element in PAP is not known. The presented results show that the template region has an essential function in both enzymes, while the second element is surprisingly dispensable in PAP. The data support the idea that the bacterial PAP descends from CCA-adding enzymes and still carries some of the structural elements required for CCA-addition as an evolutionary relic and is now fixed in a conformation specific for A-addition

7th meeting of the global arthritis research network

Last October, the 7th meeting of the Global Arthritis Research Network was held in Zurich, Switzerland. European and American experts who have made major recent contributions to molecular biology got together to provide insights into novel technologies and approaches useful for biomedical research, especially for research on arthritis and related conditions

Expression and function of EZH2 in synovial fibroblasts: epigenetic repression of the Wnt inhibitor SFRP1 in rheumatoid arthritis

OBJECTIVES: /st> To study the expression, regulation and function of the histone methyltransferase enhancer of zeste homologue 2 (EZH2) in synovial fibroblasts (SF) from patients with rheumatoid arthritis (RA) and osteoarthritis (OA). METHODS: /st> SF were obtained from RA and OA patients undergoing joint surgery. Expression levels were assessed by quantitative real-time PCR and western blot. Kinase inhibitors and reporter gene assays were employed to study signalling pathways. Functional analyses included EZH2 overexpression by plasmid transfection and gene silencing by small interfering RNA. Chromatin immunoprecipitation assay was used to analyse histone methylation within distinct promoter regions. RESULTS: /st> By studying the expression and function of EZH2 in SF the authors found that EZH2 is overexpressed in rheumatoid arthritis synovial fibroblasts (RASF) and further induced by tumour necrosis factor alpha through the nuclear factor kappa B and Jun kinase pathways. As a target gene of EZH2 the authors identified secreted frizzled-related protein 1 (SFRP1), an inhibitor of Wnt signalling, which is associated with the activation of RASF, and show that SFRP1 expression correlates with the occupation of its promoter with activating and silencing histone marks. CONCLUSIONS: /st> These data strongly suggest that the chronic inflammatory environment of the RA joint induces EZH2 and thus might cause changes in the epigenetic programmes of SF

Interleukin-6 Modulates the Expression of the Bone Morphogenic Protein Receptor Type II Through a Novel STAT3-microRNA Cluster 17/92 Pathway

Dysregulated expression of bone morphogenetic protein receptor type II (BMPR2) is a pathogenetic hallmark of pulmonary hypertension. Downregulation of BMPR2 protein but not mRNA has been observed in multiple animal models mimicking the disease, indicating a posttranscriptional mechanism of regulation. Because microRNAs (miRNAs) regulate gene expression mainly through inhibition of target gene translation, we hypothesized that miRNAs may play a role in the modulation of BMPR2. Performing a computational algorithm on the BMPR2 gene, several miRNAs encoded by the miRNA cluster 17/92 (miR-17/92) were retrieved as potential regulators. Ectopic overexpression of miR-17/92 resulted in a strong reduction of the BMPR2 protein, and a reporter gene system showed that BMPR2 is directly targeted by miR-17-5p and miR-20a. By stimulation experiments, we found that the miR-17/92 cluster is modulated by interleukin (IL)-6, a cytokine involved in the pathogenesis of pulmonary hypertension. Because IL-6 signaling is mainly mediated by STAT3 (signal transducer and activator of transcription 3), the expression of STAT3 was knocked down by small interfering RNA, which abolished the IL-6-mediated expression of miR-17/92. Consistent with these data, we found a highly conserved STAT3-binding site in the promoter region of the miR-17/92 gene (C13orf25). Promoter studies confirmed that IL-6 enhances transcription of C13orf25 through this distinct region. Finally, we showed that persistent activation of STAT3 leads to repressed protein expression of BMPR2. Taken together, we describe here a novel STAT3-miR-17/92-BMPR2 pathway, thus providing a mechanistic explanation for the loss of BMPR2 in the development of pulmonary hypertension

Investigations of solid liquid interfaces in ultra-thin liquid films via single particle tracking of silica nanoparticles

Single particle tracking with a wide field microscope is used to study the solid liquid interface between the viscous liquid tetrakis(2-ethylhexoxy)-silane and a silicon dioxide surface. Silicon dioxide nanoparticles (5 nm diameter) marked with the fluorescent dye rhodamine 6G are used as probes. The distributions of diffusion coefficients, obtained by mean squared displacements, reveal heterogeneities with at least two underlying diffusion components. Measurements on films with varying film thicknesses show that the slower component is independent of the film thickness, while the faster one increases with the film thickness. Additionally, we could show that the diffusion behavior of the particles cannot be sufficiently described by only two diffusion coefficients

Epigenetically-driven anatomical diversity of synovial fibroblasts guides joint-specific fibroblast functions

A number of human diseases, such as arthritis and atherosclerosis, include characteristic pathology in specific anatomical locations. Here we show transcriptomic differences in synovial fibroblasts from different joint locations and that HOX gene signatures reflect the joint-specific origins of mouse and human synovial fibroblasts and synovial tissues. Alongside DNA methylation and histone modifications, bromodomain and extra-terminal reader proteins regulate joint-specific HOX gene expression. Anatomical transcriptional diversity translates into joint-specific synovial fibroblast phenotypes with distinct adhesive, proliferative, chemotactic and matrix-degrading characteristics and differential responsiveness to TNF, creating a unique microenvironment in each joint. These findings indicate that local stroma might control positional disease patterns not only in arthritis but in any disease with a prominent stromal component. © The Author(s) 2017