Functional Annotation Of Mammalian Genome Research Articles

Expression profile of radiotherapy sensitive genes and tumor-associated immune cell infiltration and prognosis in multiple human cancers

Radiotherapy sensitive genes (RSGs), as new targets for cancer therapy, combined with tumor microenvironment (TME) and prognosis survival analysis, can provide a new idea for cancer therapy. In this study, RSGs were firstly obtained by searching the literature, then tumor and immune cell data from the Functional Annotation Of Mammalian genomes (FANTOM5) project were analyzed, with definition of two different expression patterns, tumor cell (TC) dominant and immune cell (IC) dominant, and the patterns were verified with RNA-seq data downloaded from the Gene Expression Omnibus (GEO) database. In addition, the correlation between the expression pattern of RSGs and immunotherapy was studied by integrating the seven cancers’ radiotherapy data from The Cancer Genome Atlas (TCGA). Result showed that the GEO datasets verification passing rate was greater than 60%. We determined the association between immune infiltration and patients' radiotherapy outcomes, and found that the infiltration levels of Head and Neck Squamous Carcinoma (HNSC) and Uterine Corpus Endometrial Carcinoma (UCEC) in IC-dominated RSGs were higher than those in TC, while the opposite phenomenon was observed in Glioblastoma Multiforme (GBM). The survival analysis of RSGs in seven cancers showed good prognosis in IC (70%) and TC (80%). These findings may help elucidate the influence of tumor-associated immune cell infiltration and prognosis on tumor radiotherapy outcome, which further identified radiosensitive biomarkers of clinical individualized radiotherapy.

Directional association test reveals high-quality putative cancer driver biomarkers including noncoding RNAs

BackgroundMost statistical methods used to identify cancer driver genes are either biased due to choice of assumed parametric models or insensitive to directional relationships important for causal inference. To overcome modeling biases and directional insensitivity, a recent statistical functional chi-squared test (FunChisq) detects directional association via model-free functional dependency. FunChisq examines patterns pointing from independent to dependent variables arising from linear, non-linear, or many-to-one functional relationships. Meanwhile, the Functional Annotation of Mammalian Genome 5 (FANTOM5) project surveyed gene expression at over 200,000 transcription start sites (TSSs) in nearly all human tissue types, primary cell types, and cancer cell lines. The data cover TSSs originated from both coding and noncoding genes. For the vast uncharacterized human TSSs that may exhibit complex patterns in cancer versus normal tissues, the model-free property of FunChisq provides us an unprecedented opportunity to assess the evidence for a gene’s directional effect on human cancer.ResultsWe first evaluated FunChisq and six other methods using 719 curated cancer genes on the FANTOM5 data. FunChisq performed best in detecting known cancer driver genes from non-cancer genes. We also show the capacity of FunChisq to reveal non-monotonic patterns of functional association, to which typical differential analysis methods such as t-test are insensitive. Further applying FunChisq to screen unannotated TSSs in FANTOM5, we predicted 1108 putative cancer driver noncoding RNAs, stronger than 90% of curated cancer driver genes. Next, we compared leukemia samples against other samples in FANTOM5 and FunChisq predicted 332/79 potential biomarkers for lymphoid/myeloid leukemia, stronger than the TSSs of all 87/100 known driver genes in lymphoid/myeloid leukemia.ConclusionsThis study demonstrated the advantage of FunChisq in revealing directional association, especially in detecting non-monotonic patterns. Here, we also provide the most comprehensive catalog of high-quality biomarkers that may play a causative role in human cancers, including putative cancer driver noncoding RNAs and lymphoid/myeloid leukemia specific biomarkers.

An integrated expression atlas of miRNAs and their promoters in human and mouse.

MicroRNAs (miRNAs) are short non-coding RNAs with key roles in cellular regulation. As part of the fifth edition of the Functional Annotation of Mammalian Genome (FANTOM5) project, we created an integrated expression atlas of miRNAs and their promoters by deep-sequencing 492 short RNA (sRNA) libraries, with matching Cap Analysis Gene Expression (CAGE) data, from 396 human and 47 mouse RNA samples. Promoters were identified for 1,357 human and 804 mouse miRNAs and showed strong sequence conservation between species. We also found that primary and mature miRNA expression levels were correlated, allowing us to use the primary miRNA measurements as a proxy for mature miRNA levels in a total of 1,829 human and 1,029 mouse CAGE libraries. We thus provide a broad atlas of miRNA expression and promoters in primary mammalian cells, establishing a foundation for detailed analysis of miRNA expression patterns and transcriptional control regions.

Expression Specificity of Disease-Associated lncRNAs: Toward Personalized Medicine.

Long noncoding RNAs (lncRNAs) perform diverse regulatory functions in transcription, translation' chromatin modification, and cellular organization. Misregulation of lncRNAs is found linked to various human diseases. Compared to protein-coding RNAs' lncRNAs are more specific to organs, tissues, cell types, developmental stages, and disease conditions' making them promising candidates as diagnostic and prognostic biomarkers and as gene therapy targets. The functional annotation of mammalian genome (FANTOM) consortium utilizes cap analysis of gene expression (CAGE) method to quantify genome-wide activities of promoters and enhancers of coding and noncoding RNAs across a large collection of human and mouse tissues' cell types' diseases, and time-courses. The project discovered widespread transcription of major lncRNA classes, including lncRNAs derived from enhancers' bidirectional promoters' antisense lncRNAs' and repetitive elements. Results from FANTOM project enable assessment of lncRNA expression specificity across tissue and disease conditions' based on differential promoter and enhancer usage. More than 85% of disease-related SNPs are within noncoding regions and are strikingly overrepresented in enhancer and promoter regions, suggestive of the importance of lncRNA loci at these SNP harboring regions to human diseases. In this chapter' we discuss lncRNA expression specificity' review diverse functions of disease-associated lncRNAs' and present perspectives on their potential therapeutic applications for personalized medicine. The future development of lncRNA applications relies on technologies to identify and validate their functions' structures' and mechanisms. Comprehensive understanding of genome-wide interaction networks of lncRNAs with proteins, chromatins, and other RNAs in regulating cellular processes will allow personalized medicine to use lncRNAs as highly specific biomarkers in diagnosis' prognosis, and therapeutic targets.

Macrophages.com: An on-line community resource for innate immunity research

Macrophages play a major role in tissue remodelling during development, wound healing and tissue homeostasis, and are central to innate immunity and to the pathology of tissue injury and inflammation. Given this fundamental role in many aspects of biological function, an enormous wealth of information has accumulated on these fascinating cells in the literature and other public repositories. With the escalation of genome-scale data derived from macrophages and related haematopoietic cell types, there is a growing need for an integrated resource that seeks to compile, organise and analyse our collective knowledge of macrophage biology. Here we describe a community-driven web-based resource, macrophages.com that aims to provide a portal onto various types of Omics data to facilitate comparative genomic studies, promoter and transcriptional network analyses, models of macrophage pathways together with other information on these cells. To this end, the website combines public and in-house analyses of expression data with pre-analysed views of co-expressed genes as supported by the network analysis tool BioLayout Express 3D, as well as providing access to maps of pathways active in macrophages. Macrophages.com also provides access to an extensive image library of macrophages in adult/embryonic tissue sections prepared from normal and transgenic mice. In addition, the site links to the Human Protein Atlas database so as to provide direct access to protein expression patterns in human macrophages. Finally, an integrated gene-centric portal provides the tools for rapid promoter analysis studies based on a comprehensive set of CAGE-derived transcription start site (TSS) sequences in human and mouse genomes as generated by the Functional Annotation of Mammalian genomes (FANTOM) projects initiated by the RIKEN Omics Science Center. Our aim is to continue to grow the macrophages.com resource using publicly available data, as well as in-house generated knowledge. In so doing we aim to provide a user-friendly community website and a community portal for access to comprehensive sets of macrophage-related data.

Beyond the FANTOM4

RIKEN OSC has a long history of transcriptome analysis using DNA sequencers; we also organize the international consortium Functional Annotation of Mammalian Genome (FANTOM; Figure ​Figure1).1). We have developed a novel method Cap Analysis of Gene Expression (CAGE), which analyzes capped transcripts in a genome-wide manner by sequencing the 5' end of mRNA. In the recently completed FANTOM4, we analyzed the transcriptional regulation network of human THP-1 cell differentiation at the promoter level with deepCAGE, an application of CAGE for the next-generation sequencers. The deepCAGE data were also used to analyze the promoter activity of repeat elements and to discover new small RNA (tiRNA) by integration with the small RNA sequencing. Figure 1 History of FANTOMs. Although the deepCAGE is a powerful tool for transcriptome analysis, it is limited owing to the need to use a significantly large amount of RNA and it is semi-quantitative. We have addressed these problems and developed NanoCAGE technology and quantitative deepCAGE. OSC aims to expand transcriptome knowledge; we aim to understand promoter and transcriptional regulation. We are about to start the FANTOM5 project.

A single lentiviral vector platform for microRNA-based conditional RNA interference and coordinated transgene expression.

RNAi is proving to be a powerful experimental tool for the functional annotation of mammalian genomes. The full potential of this technology will be realized through development of approaches permitting regulated manipulation of endogenous gene expression with coordinated reexpression of exogenous transgenes. We describe the development of a lentiviral vector platform, pSLIK (single lentivector for inducible knockdown), which permits tetracycline-regulated expression of microRNA-like short hairpin RNAs from a single viral infection of any naïve cell system. In mouse embryonic fibroblasts, the pSLIK platform was used to conditionally deplete the expression of the heterotrimeric G proteins Galpha12 and Galpha13 both singly and in combination, demonstrating the Galpha13 dependence of serum response element-mediated transcription. In RAW264.7 macrophages, regulated knockdown of Gbeta2 correlated with a reduced Ca(2+) response to C5a. Insertion of a GFP transgene upstream of the Gbeta2 microRNA-like short hairpin RNA allowed concomitant reexpression of a heterologous mRNA during tetracycline-dependent target gene knockdown, significantly enhancing the experimental applicability of the pSLIK system.

In vivo functional proteomics: mammalian genome annotation using CD-tagging.

A self-inactivating CD-tagging retroviral vector was used to introduce epitope and GFP tags into genes and proteins in NIH 3T3 cells. Several hundred cell clones, each expressing GFP fluorescence in a distinctive pattern, were isolated. Molecular analysis showed that a wide variety of genes and proteins, some known and some newly discovered, had been tagged. The analysis also revealed that, in the great majority of instances, the abundance and cellular location of the tagged protein mirrored that of its untagged counterpart. This approach provides a systematic means for the functional annotation of mammalian genomes and proteomes in living cells.

Engineering chromosomal rearrangements in mice.

The combination of gene-targeting techniques in mouse embryonic stem cells and the Cre/loxP site-specific recombination system has resulted in the emergence of chromosomal-engineering technology in mice. This advance has opened up new opportunities for modelling human diseases that are associated with chromosomal rearrangements. It has also led to the generation of visibly marked deletions and balancer chromosomes in mice, which provide essential reagents for maximizing the efficiency of large-scale mutagenesis efforts and which will accelerate the functional annotation of mammalian genomes, including the human genome.