The Two Stem Cell MicroRNA Gene Clusters C19MC and miR-371-3 Are Activated by Specific Chromosomal Rearrangements in a Subgroup of Thyroid Adenomas

Thyroid adenomas are common benign human tumors with a high prevalence of about 5% of the adult population even in iodine sufficient areas. Rearrangements of chromosomal band 19q13.4 represent a frequent clonal cytogenetic deviation in these tumors making them the most frequent non-random chromosomal translocations in human epithelial tumors at all. Two microRNA (miRNA) gene clusters i.e. C19MC and miR-371-3 are located in close proximity to the breakpoint region of these chromosomal rearrangements and have been checked for a possible up-regulation due to the genomic alteration. In 4/5 cell lines established from thyroid adenomas with 19q13.4 rearrangements and 5/5 primary adenomas with that type of rearrangement both the C19MC and miR-371-3 cluster were found to be significantly overexpressed compared to controls lacking that particular chromosome abnormality. In the remaining cell line qRT-PCR revealed overexpression of members of the miR-371-3 cluster only which might be due to a deletion accompanying the chromosomal rearrangement in that case. In depth molecular characterization of the breakpoint in a cell line from one adenoma of this type reveals the existence of large Pol-II mRNA fragments as the most likely source of up-regulation of the C19MC cluster. The up-regulation of the clusters is likely to be causally associated with the pathogenesis of the corresponding tumors. Of note, the expression of miRNAs miR-520c and miR-373 is known to characterize stem cells and in terms of molecular oncology has been implicated in invasive growth of epithelial cells in vitro and in vivo thus allowing to delineate a distinct molecular subtype of thyroid adenomas. Besides thyroid adenomas rearrangements of 19q13.4 are frequently found in other human neoplasias as well, suggesting that activation of both clusters might be a more general phenomenon in human neoplasias

Molecular topography of the MED12-deleted region in smooth muscle tumors: a possible link between non-B DNA structures and hypermutability

Abstract Background Deletions of the gene encoding mediator subcomplex 12 (MED12) in human smooth muscle tumors rank among the most frequent genomic alterations in human tumors at all. In a minority of these cases, small deletions are found. In an attempt to delineate key features of the deletions aimed at a better understanding of the molecular pathogenesis of uterine smooth muscle tumors we have analyzed 70 MED12 deletions including 46 cases from the literature and 24 own unpublished cases. Results The average length of the deletions was 18.7 bp ranging between 2 bp and 43 bp. While in general multitudes of 3 clearly dominated leaving the transcript in frame, deletions of 21, 24, 30, and 33 nucleotides were clearly underrepresented. Within the DNA segment affected deletion breakpoints were not randomly distributed. Most breakpoints clustered within the center of the segment where two peaks of breakpoint clusters could be distinguished. Interestingly, one of these clusters coincides with the loop of a putative folded non-B DNA structure whereas a much lower number of breaks noted in the 5′ and 3′ stem of the structure forming an intramolecular B-helix. The second cluster mainly consisting of 3′ breaks was located in a region downstream adjacent to the stem. Conclusion The present study describes for the first time main characteristics of MED12 deletions occurring in smooth muscle tumors. Interestingly, the non-random distribution of breakpoints within the deletion hotspot region may point to a role of non-canonical DNA structures for the occurrence of these mutations and the molecular pathogenesis of uterine smooth muscle tumors, respectively. </jats:sec

Crystalline iron oxides stimulate methanogenic benzoate degradation in marine sediment- derived enrichment cultures

Elevated dissolved iron concentrations in the methanic zone are typical geochemical signatures of rapidly accumulating marine sediments. These sediments are often characterized by co-burial of iron oxides with recalcitrant aromatic organic matter of terrigenous origin. Thus far, iron oxides are predicted to either impede organic matter degradation, aiding its preservation, or identified to enhance organic carbon oxidation via direct electron transfer. Here, we investigated the effect of various iron oxide phases with differing crystallinity (magnetite, hematite, and lepidocrocite) during microbial degradation of the aromatic model compound benzoate in methanic sediments. In slurry incubations with magnetite or hematite, concurrent iron reduction, and methanogenesis were stimulated during accelerated benzoate degradation with methanogenesis as the dominant electron sink. In contrast, with lepidocrocite, benzoate degradation, and methanogenesis were inhibited. These observations were reproducible in sediment-free enrichments, even after five successive transfers. Genes involved in the complete degradation of benzoate were identified in multiple metagenome assembled genomes. Four previously unknown benzoate degraders of the genera Thermincola (Peptococcaceae, Firmicutes), Dethiobacter (Syntrophomonadaceae, Firmicutes), Deltaproteobacteria bacteria SG8_13 (Desulfosarcinaceae, Deltaproteobacteria), and Melioribacter (Melioribacteraceae, Chlorobi) were identified from the marine sediment-derived enrichments. Scanning electron microscopy (SEM) and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) images showed the ability of microorganisms to colonize and concurrently reduce magnetite likely stimulated by the observed methanogenic benzoate degradation. These findings explain the possible contribution of organoclastic reduction of iron oxides to the elevated dissolved Fe2+ pool typically observed in methanic zones of rapidly accumulating coastal and continental margin sediments

Chromatin Immunoprecipitation to Analyze DNA Binding Sites of HMGA2

BACKGROUND: HMGA2 is an architectonic transcription factor abundantly expressed during embryonic and fetal development and it is associated with the progression of malignant tumors. The protein harbours three basically charged DNA binding domains and an acidic protein binding C-terminal domain. DNA binding induces changes of DNA conformation and hence results in global overall change of gene expression patterns. Recently, using a PCR-based SELEX (Systematic Evolution of Ligands by Exponential Enrichment) procedure two consensus sequences for HMGA2 binding have been identified. METHODOLOGY/PRINCIPAL FINDINGS: In this investigation chromatin immunoprecipitation (ChIP) experiments and bioinformatic methods were used to analyze if these binding sequences can be verified on chromatin of living cells as well. CONCLUSION: After quantification of HMGA2 protein in different cell lines the colon cancer derived cell line HCT116 was chosen for further ChIP experiments because of its 3.4-fold higher HMGA2 protein level. 49 DNA fragments were obtained by ChIP. These fragments containing HMGA2 binding sites have been analyzed for their AT-content, location in the human genome and similarities to sequences generated by a SELEX study. The sequences show a significantly higher AT-content than the average of the human genome. The artificially generated SELEX sequences and short BLAST alignments (11 and 12 bp) of the ChIP fragments from living cells show similarities in their organization. The flanking regions are AT-rich, whereas a lower conservation is present in the center of the sequences

Decrease in thyroid adenoma associated (THADA) expression is a marker of dedifferentiation of thyroid tissue

<p>Abstract</p> <p>Background</p> <p><it>Thyroid adenoma associated (THADA) </it>has been identified as the target gene affected by chromosome 2p21 translocations in thyroid adenomas, but the role of THADA in the thyroid is still elusive. The aim of this study was to quantify <it>THADA </it>gene expression in normal tissues and in thyroid hyper- and neoplasias, using real-time PCR.</p> <p>Methods</p> <p>For the analysis <it>THADA </it>and 18S rRNA gene expression assays were performed on 34 normal tissue samples, including thyroid, salivary gland, heart, endometrium, myometrium, lung, blood, and adipose tissue as well as on 85 thyroid hyper- and neoplasias, including three adenomas with a 2p21 translocation. In addition, <it>NIS </it>(<it>sodium-iodide symporter</it>) gene expression was measured on 34 of the pathological thyroid samples.</p> <p>Results</p> <p>Results illustrated that <it>THADA </it>expression in normal thyroid tissue was significantly higher (<it>p </it>< 0.0001, exact Wilcoxon test) than in the other tissues. Significant differences were also found between non-malignant pathological thyroid samples (goiters and adenomas) and malignant tumors (<it>p </it>< 0.001, Wilcoxon test, t approximation), anaplastic carcinomas (ATCs) and all other samples and also between ATCs and all other malignant tumors (<it>p </it>< 0.05, Wilcoxon test, t approximation). Furthermore, in thyroid tumors <it>THADA </it>mRNA expression was found to be inversely correlated with <it>HMGA2 </it>mRNA. <it>HMGA2 </it>expression was recently identified as a marker revealing malignant transformation of thyroid follicular tumors. A correlation between <it>THADA </it>and <it>NIS </it>has also been found in thyroid normal tissue and malignant tumors.</p> <p>Conclusions</p> <p>The results suggest <it>THADA </it>being a marker of dedifferentiation of thyroid tissue.</p

Causal discovery of gene regulation with incomplete data

Causal discovery algorithms aim to identify causal relations from observational data and have become a popular tool for analysing genetic regulatory systems. In this work, we applied causal discovery to obtain novel insights into the genetic regulation underlying head‐and‐neck squamous cell carcinoma. Some methodological challenges needed to be resolved first. The available data contained missing values, but most approaches to causal discovery require complete data. Hence, we propose a new procedure combining constraint‐based causal discovery with multiple imputation. This is based on using Rubin's rules for pooling tests of conditional independence. A second challenge was that causal discovery relies on strong assumptions and can be rather unstable. To assess the robustness of our results, we supplemented our investigation with sensitivity analyses, including a non‐parametric bootstrap to quantify the variability of the estimated causal structures. We applied these methods to investigate how the high mobility group AT‐Hook 2 (HMGA2) gene is incorporated in the protein 53 signalling pathway playing an important role in head‐and‐neck squamous cell carcinoma. Our results were quite stable and found direct associations between HMGA2 and other relevant proteins, but they did not provide clear support for the claim that HMGA2 itself is a key regulator gene

Effects of oxygen availability on mycobenthic communities of marine coastal sediments

Abstract In coastal marine sediments, oxygen availability varies greatly, and anoxic conditions can develop quickly over low spatial resolution. Although benthic fungi are important players in the marine carbon cycle, little is known about their adaptation to fluctuating availability of oxygen as terminal electron acceptor. Here, we study which part of a mycobenthic community from oxic coastal sediments can thrive under temporarily anoxic conditions. We test whether phylogeny or certain fungal traits promote plasticity in respect to changes in oxygen availability. Therefore, we incubated mycobenthos under oxic and anoxic conditions, performed ITS2 Illumina tag-sequencing and an additional meta-analysis on a literature survey. Half of all OTUs showed a plasticity towards changing oxygen availability and exhibited different strategies towards anoxic conditions, with rapid response within hours or a delayed one after several days. The strategy of dimorphism and facultative yeasts were significantly linked to OTU occurrence in anoxic conditions, while phylogeny and other traits had less effect. Our results suggest that different fungal niches are formed over the duration of prolonged anoxic conditions. The taxon-specific proliferation seems to be regulated by the fine-tuning of various traits and factors. It is essential to take these results into account when conducting conceptual work on the functionality of the marine benthos

Causal Discovery of Gene Regulation with Incomplete Data

Summary Causal discovery algorithms aim to identify causal relations from observational data and have become a popular tool for analysing genetic regulatory systems. In this work, we applied causal discovery to obtain novel insights into the genetic regulation underlying head-and-neck squamous cell carcinoma. Some methodological challenges needed to be resolved first. The available data contained missing values, but most approaches to causal discovery require complete data. Hence, we propose a new procedure combining constraint-based causal discovery with multiple imputation. This is based on using Rubin's rules for pooling tests of conditional independence. A second challenge was that causal discovery relies on strong assumptions and can be rather unstable. To assess the robustness of our results, we supplemented our investigation with sensitivity analyses, including a non-parametric bootstrap to quantify the variability of the estimated causal structures. We applied these methods to investigate how the high mobility group AT-Hook 2 (HMGA2) gene is incorporated in the protein 53 signalling pathway playing an important role in head-and-neck squamous cell carcinoma. Our results were quite stable and found direct associations between HMGA2 and other relevant proteins, but they did not provide clear support for the claim that HMGA2 itself is a key regulator gene.</jats:p