Genome-wide perturbations of Alu expression and Alu-associated post-transcriptional regulations distinguish oligodendroglioma from other gliomas.

Alu is a primate-specific repeat element in the human genome and has been increasingly appreciated as a regulatory element in many biological processes. But the appreciation of Alu has been limited in tumorigenesis, especially for brain tumor. To investigate the relevance of Alu to the gliomagenesis, we studied Alu element-associated post-transcriptional processes and the RNA expression of the element by performing RNA-seq for a total of 41 pairs of neurotypical and diverse glioma brain tissues. We find that A-to-I editing and circular RNA levels, as well as Alu RNA expression, are decreased overall in gliomas, compared to normal tissue. Interestingly, grade 2 oligodendrogliomas are least affected in A-to-I editing and circular RNA levels among gliomas, whereas they have a higher proportion of down-regulated Alu subfamilies, compared to the other gliomas. These findings collectively imply a unique pattern of Alu-associated transcriptomes in grade 2 oligodendroglioma, providing an insight to gliomagenesis from the perspective of an evolutionary genetic element.

The Firre locus produces a trans-acting RNA molecule that functions in hematopoiesis.

RNA has been classically known to play central roles in biology, including maintaining telomeres, protein synthesis, and in sex chromosome compensation. While thousands of long noncoding RNAs (lncRNAs) have been identified, attributing RNA-based roles to lncRNA loci requires assessing whether phenotype(s) could be due to DNA regulatory elements, transcription, or the lncRNA. Here, we use the conserved X chromosome lncRNA locus Firre, as a model to discriminate between DNA- and RNA-mediated effects in vivo. We demonstrate that (i) Firre mutant mice have cell-specific hematopoietic phenotypes, and (ii) upon exposure to lipopolysaccharide, mice overexpressing Firre exhibit increased levels of pro-inflammatory cytokines and impaired survival. (iii) Deletion of Firre does not result in changes in local gene expression, but rather in changes on autosomes that can be rescued by expression of transgenic Firre RNA. Together, our results provide genetic evidence that the Firre locus produces a trans-acting lncRNA that has physiological roles in hematopoiesis.

Genome-wide investigation of intragenic DNA methylation identifies ZMIZ1 gene as a prognostic marker in glioblastoma and multiple cancer types.

DNA methylation has long been recognized as a tumor-promoting factor when aberrantly regulated in the promoter region of genes. However, the effect of intragenic DNA methylation remains poorly understood on the clinical aspects of cancer. Here, we first evaluated the significance of intragenic DNA methylation for survival outcomes of cancer patients in a genome-wide manner. Glioblastoma patients with hypermethylated intragenic regions exhibited better survival than hypomethylated patients. Enrichment analyses of intragenic DNA methylation profiles with epigenetic signatures prioritized the intragenic DNA methylation of ZMIZ1 as a possible glioblastoma prognostic marker that is independent of MGMT methylation in IDH1 wild-type patients. This intragenic region harbored molecular signatures of alternative transcription across many cell types. Furthermore, we found that the intragenic region of ZMIZ1 can serve as a molecular marker in multiple cancers including astrocytomas, bladder cancer and renal cell carcinoma according to DNA methylation status. Finally, in vitro and in vivo experiments uncovered the role of ZMIZ1 as a driver of tumor cell migration. Altogether, our results identify ZMIZ1 as a prognostic marker in cancer and highlight the clinical significance of intragenic methylation in cancer.

Integrative analysis of DNA methylation suggests down-regulation of oncogenic pathways and reduced somatic mutation rates in survival outliers of glioblastoma.

The study of survival outliers of glioblastoma can provide important clues on gliomagenesis as well as on the ways to alter clinical course of this almost uniformly lethal cancer type. However, there has been little consensus on genetic and epigenetic signatures of the long-term survival outliers of glioblastoma. Although the two classical molecular markers of glioblastoma including isocitrate dehydrogenase 1 or 2 (IDH1/2) mutation and O6-methylguanine DNA methyltransferase (MGMT) promoter methylation are associated with overall survival rate of glioblastoma patients, they are not specific to the survival outliers. In this study, we compared the two groups of survival outliers of glioblastoma with IDH wild-type, consisting of the glioblastoma patients who lived longer than 3 years (n = 17) and the patients who lived less than 1 year (n = 12) in terms of genome-wide DNA methylation profile. Statistical analyses were performed to identify differentially methylated sites between the two groups. Functional implication of DNA methylation patterns specific to long-term survivors of glioblastoma were investigated by comprehensive enrichment analyses with genomic and epigenomic features. We found that the genome of long-term survivors of glioblastoma is differentially methylated relative to short-term survivor patients depending on CpG density: hypermethylation near CpG islands (CGIs) and hypomethylation far from CGIs. Interestingly, these two patterns are associated with distinct oncogenic aspects in gliomagenesis. In the long-term survival glioblastoma-specific sites distant from CGI, somatic mutations of glioblastoma are enriched with higher DNA methylation, suggesting that the hypomethylation in long-term survival glioblastoma can contribute to reduce the rate of somatic mutation. On the other hand, the hypermethylation near CGIs associates with transcriptional downregulation of genes involved in cancer progression pathways. Using independent cohorts of IDH1/2- wild type glioblastoma, we also showed that these two patterns of DNA methylation can be used as molecular markers of long-term survival glioblastoma. Our results provide extended understanding of DNA methylation, especially of DNA hypomethylation, in cancer genome and reveal clinical importance of DNA methylation pattern as prognostic markers of glioblastoma.

Influence of heat and moisture treatment on carotenoids, phenolic content, and antioxidant capacity of orange maize flour.

The aim of this work was to study the effect of heat and moisture treatment (HMT) on carotenoids, phenolic content and antioxidant capacity of ground, orange maize. Total carotenoid content (TCC) of untreated sample (53.39 mg/kg) was 2.2 times higher than measured in treated orange maize f (24.61 mg/kg). The rates of degradation with HMT were in the following order: ß-carotene > ß-cryptoxanthin > zeaxanthin > lutein. There was a significant interaction between longer heating time and higher moisture content on carotenoid degradation (p < .05). Total phenolic content (TPC) in raw sample (1664.74 mg/kg) was two-fold higher than in treated orange maize (827.89 mg/kg). Ferulic acid was the most abundant and stable phenolic acid in raw and treated orange maize. The antioxidant capacity of orange maize was higher in methanol than in butanol extracts. The highest correlation (0.924) was observed between TPC and ABTS+ scavenging capacity of methanol extracts.

Restoration of miR-29b exerts anti-cancer effects on glioblastoma.

BACKGROUND: Glioblastoma multiforme (GBM) is known as one of the most fatal forms of cancer. MicroRNAs have been widely implicated in the regulation of mammalian development and pathogenesis. The brain-enriched miR-29 subfamilies are known to be exclusively expressed in the developing brain, and they are aberrantly down-regulated in GBM. This study aims to elucidate the role of miR-29b in GBM development and the feasibility of therapeutic targeting using conjugated nanoparticles. METHODS: After confirmation of miR-29b expression levels in GBM tissues by analysis of open source data, the anticancer effect of miR-29b was tested by the introduction of syn-hsa-miR-29b-3p in the A172 GBM cell line. In vitro studies of cell viability and apoptosis and ex vivo study using GBM tissue slice cultures from 3 patients and nanoparticle delivery of miR-29b were performed. RESULTS: We discovered an increase in apoptotic cell populations with the introduction of miR-29b in the GBM cell line. An established human-derived GBM tissue slice culture system confirmed the anticancer effect of miR-29b-conjugated nanoparticles. Using PCR array, we found that exogenous miR-29b inhibits the expression of COL1A2, COL3A1, COL4A1, ELN, ITGA11, MMP24, and SPARC, which mediates an anticancer effect. CONCLUSIONS: miR-29b may serve as a putative therapeutic molecule when its expression is restored using a nanoparticle delivery system in GBM.

Glutaminase 2 expression is associated with regional heterogeneity of 5-aminolevulinic acid fluorescence in glioblastoma.

Fluorescence-guided surgery using 5-aminolevulinic acid (5-ALA) is now a widely-used modality for glioblastoma (GBM) treatment. However, intratumoral heterogeneity of fluorescence intensity may reflect different onco-metabolic programs. Here, we investigated the metabolic mechanism underlying the heterogeneity of 5-ALA fluorescence in GBM. Using an in-house developed fluorescence quantification system for tumor tissues, we collected 3 types of GBM tissues on the basis of their fluorescence intensity, which was characterized as strong, weak, and none. Expression profiling by RNA-sequencing revealed 77 genes with a proportional relationship and 509 genes with an inverse relationship between gene expression and fluorescence intensity. Functional analysis and in vitro experiments confirmed glutaminase 2 (GLS2) as a key gene associated with the fluorescence heterogeneity. Subsequent metabolite profiling discovered that insufficient NADPH due to GLS2 underexpression was responsible for the delayed metabolism of 5-ALA and accumulation of protoporphyrin IX (PpIX) in the high fluorescence area. The expression level of GLS2 and related NADPH production capacity is associated with the regional heterogeneity of 5-ALA fluorescence in GBM.

Dynamic regulation of RNA editing in human brain development and disease.

RNA editing is increasingly recognized as a molecular mechanism regulating RNA activity and recoding proteins. Here we surveyed the global landscape of RNA editing in human brain tissues and identified three unique patterns of A-to-I RNA editing rates during cortical development: stable high, stable low and increasing. RNA secondary structure and the temporal expression of adenosine deaminase acting on RNA (ADAR) contribute to cis- and trans-regulatory mechanisms of these RNA editing patterns, respectively. Interestingly, the increasing pattern was associated with neuronal maturation, correlated with mRNA abundance and potentially influenced miRNA binding energy. Gene ontology analyses implicated the increasing pattern in vesicle or organelle membrane-related genes and glutamate signaling pathways. We also found that the increasing pattern was selectively perturbed in spinal cord injury and glioblastoma. Our findings reveal global and dynamic aspects of RNA editing in brain, providing new insight into epitranscriptional regulation of sequence diversity.

Identification of differentially expressed subnetworks based on multivariate ANOVA.

BACKGROUND: Since high-throughput protein-protein interaction (PPI) data has recently become available for humans, there has been a growing interest in combining PPI data with other genome-wide data. In particular, the identification of phenotype-related PPI subnetworks using gene expression data has been of great concern. Successful integration for the identification of significant subnetworks requires the use of a search algorithm with a proper scoring method. Here we propose a multivariate analysis of variance (MANOVA)-based scoring method with a greedy search for identifying differentially expressed PPI subnetworks. RESULTS: Given the MANOVA-based scoring method, we performed a greedy search to identify the subnetworks with the maximum scores in the PPI network. Our approach was successfully applied to human microarray datasets. Each identified subnetwork was annotated with the Gene Ontology (GO) term, resulting in the phenotype-related functional pathway or complex. We also compared these results with those of other scoring methods such as t statistic- and mutual information-based scoring methods. The MANOVA-based method produced subnetworks with a larger number of proteins than the other methods. Furthermore, the subnetworks identified by the MANOVA-based method tended to consist of highly correlated proteins. CONCLUSION: This article proposes a MANOVA-based scoring method to combine PPI data with expression data using a greedy search. This method is recommended for the highly sensitive detection of large subnetworks.

Structure, expression, and mapping of two nodule-specific genes identified by mining public soybean EST databases.

Numerous nodule-specific genes, which are involved in the root nodule development and function, have been known and are still being discovered. Here, we reported the structure, expression, and genetic map location of two novel nodule-specific genes. First, two EST groups, one obtained from a nodule library and the other from all aboveground tissue libraries, were clustered with regard to in silico expression profiles. We compiled a pool of 103 putative nodule-specific sequence clusters. Then, two representative ESTs were selected for further experimental analyses. According to the full-length cDNA sequences, one was an EST of a novel nodule-specific polygalacturonase gene, GmPGN, and the other was an EST of a new short nodule-specific gene, GmEKN. The results of expression analyses of the GmPGN cDNAs indicated that GmPGN expression was not detectable in any of the soybean tissues except in the nodule tissue and may be regulated via alternative splicing. GmEKN expression was the most strongly detected in the nodule. The predicted GmEKN protein is both glutamic acid- and lysine-rich, and is also highly hydrophilic. Genetic mapping located GmPGN near the known quantitative trait locus conferring resistance to soybean cyst nematode on soybean molecular linkage group (MLG) B1, and GmEKN on MLG A2. These results provide useful information for the use of these genes in research on the orchestration of numerous genes in nodule development and function.