Mouse Genome Research Articles

Prediction and comparative analysis of CTCF binding sites based on a first principle approach

We calculated the patterns for the CCCTC transcription factor (CTCF) binding sites across many genomes on a first principle approach. The validation of the first principle method was done on the human as well as on the mouse genome. The predicted human CTCF binding sites are consistent with the consensus sequence, ChIP-seq data for the K562 cell, nucleosome positions for IMR90 cell as well as the CTCF binding sites in the mouse HOXA gene. The analysis of Homo sapiens, Mus musculus, Sus scrofa, Capra hircus and Drosophila melanogaster whole genomes shows: binding sites are organized in cluster-like groups, where two consecutive sites obey a power-law with coefficient ranging from 0.3292 ± 0.0068 to 0.5409 ± 0.0064; the distance between these groups varies from 18.08 ± 0.52 kbp to 42.1 ± 2.0 kbp. The genome of Aedes aegypti does not show a power law, but 19.9% of binding sites are 144 ± 4 and 287 ± 5 bp distant of each other. We run negative tests, confirming the under-representation of CTCF binding sites in Caenorhabditis elegans, Plasmodium falciparum and Arabidopsis thaliana complete genomes.

Identification of the Transgene Integration Site and Host Genome Changes in MRP8-Cre/ires-EGFP Transgenic Mice by Targeted Locus Amplification.

The MRP8-Cre-ires/EGFP transgenic mouse (Mrp8creTg, on C57BL/6J genetic background) is popular in immunological and hematological research for specifically expressing Cre recombinase and an EGFP reporter in neutrophils. It is often crossed with other transgenic lines carrying loxP-flanked genes to achieve restricted gene knockout in neutrophils. However, due to the way in which the line was created, basic knowledge about the MRP8-Cre-ires/EGFP transgene in the host genome, such as its integration site(s) and flanking sequences, remains largely unknown, hampering robust experimental design and data interpretation. Here we used a recently developed technique, targeted locus amplification (TLA) sequencing, to fill these knowledge gaps. We found that the MRP8-Cre-ires/EGFP transgene was integrated into chromosome 5 (5qG2) of the host mouse genome. This integration led to a 44 kb deletion of the host genomic sequence, resulting in complete deletion of Serpine1 and partial deletion of Ap1s1. Having determined the flanking sequences of the transgene, we designed a new genotyping protocol that can distinguish homozygous, heterozygous, and wildtype Mrp8creTg mice. To our surprise, crossing heterozygous mice produced no homozygous Mrp8creTg mice, most likely due to prenatal lethality resulting from disrupted Ap1s1 gene expression.

A versatile transposon-based technology to generate loss- and gain-of-function phenotypes in the mouse liver

BackgroundUnderstanding the contribution of gene function in distinct organ systems to the pathogenesis of human diseases in biomedical research requires modifying gene expression through the generation of gain- and loss-of-function phenotypes in model organisms, for instance, the mouse. However, methods to modify both germline and somatic genomes have important limitations that prevent easy, strong, and stable expression of transgenes. For instance, while the liver is remarkably easy to target, nucleic acids introduced to modify the genome of hepatocytes are rapidly lost, or the transgene expression they mediate becomes inhibited due to the action of effector pathways for the elimination of exogenous DNA. Novel methods are required to overcome these challenges, and here we develop a somatic gene delivery technology enabling long-lasting high-level transgene expression in the entire hepatocyte population of mice.ResultsWe exploit the fumarylacetoacetate hydrolase (Fah) gene correction-induced regeneration in Fah-deficient livers, to demonstrate that such approach stabilizes luciferase expression more than 5000-fold above the level detected in WT animals, following plasmid DNA introduction complemented by transposon-mediated chromosomal gene transfer. Building on this advancement, we created a versatile technology platform for performing gene function analysis in vivo in the mouse liver. Our technology allows the tag-free expression of proteins of interest and silencing of any arbitrary gene in the mouse genome. This was achieved by applying the HADHA/B endogenous bidirectional promoter capable of driving well-balanced bidirectional expression and by optimizing in vivo intronic artificial microRNA-based gene silencing. We demonstrated the particular usefulness of the technology in cancer research by creating a p53-silenced and hRas G12V-overexpressing tumor model.ConclusionsWe developed a versatile technology platform for in vivo somatic genome editing in the mouse liver, which meets multiple requirements for long-lasting high-level transgene expression. We believe that this technology will contribute to the development of a more accurate new generation of tools for gene function analysis in mice.

EP News: Basic and Translational

van Ouwerkerk et al (Circulation February 3, 2022; https://doi.org/10.1161/CIRCULATIONAHA.121.054347, PMID 35113653) investigated the pathogenic missense variant p.G125R in the TBX5 transcription factor that causes Holt-Oram syndrome (HOS; hand-heart syndrome) and early onset of atrial fibrillation (AF). The TBX5-p.G125R variant was introduced in the mouse genome (Tbx5G125R). The authors observed a high incidence of atrial extrasystoles and atrioventricular conduction disturbances in patients with HOS.

Genome-wide identification, characterization, and expression analysis of keratin genes (KRTs) family in yak (Bos grunniens).

As the largest subgroup of intermediate filament proteins, keratins are divided into two types of subfamily. Currently, the molecular mechanism of keratins in several animals has been reported but is limited in yak. Here, 53 different kinds of keratins were identified in the yak genome, including 23 type I and 30 type II keratins. Bioinformatics analysis in this study revealed that multiple phosphorylation sites were identified among all the family members. And the subcellular localization of these proteins was predicted to be in the nucleus, cytoskeleton, and cytoplasm. All keratin family proteins were unstable and the scores of instability coefficient were higher than 40. Phylogenetic analysis showed that high consistency results of the sequence conservation and grouping were found in the genomes of yak, sheep, cattle, mouse, rat, and human. Based on the expression patterns obtained from the transcriptome data, keratin genes (KRTs) were grouped into five clusters, and results also showed that KRTs were highly activated in skin tissues during the hair cycle in yak. Among the five clusters, Cluster II contained the most KRTs, which was the main expression pattern of the yak hair follicle cycle, followed by Cluster III. These results indicated the transition period from telogen to anagen and catagen to telogen were highly dynamic in yak. Gene expression correlation analysis showed that KRTs exhibited a strong correlation (mainly positive correlation) throughout the hair follicle development cycle. And the identification of hub KRTs in specific modules related to hair follicle development in this study was performed using the Weight Gene Co-Expression Network Analysis (WGCNA). Specific modules that include KRTs were darkgreen (KRT40), darkgrey (KRT5), turquoise (KRT1, KRT2, KRT10), bisque4 (KRT4), thistle2 (KRT9, KRT39), and yellowgreen (KRT24). The interaction network showed that these genes were found to be related to the regulation of cell cycle, melanogenesis, hair follicle development, keratinocyte proliferation. Our study provides theoretical support for the study of the evolutionary relationship and molecular mechanism of keratin family in B. grunnien.

Multiplexing: An Efficient Way of Genetic Monitoring of Laboratory Mice using Microsatellite Markers

Background: Inbred strains of laboratory mice have been widely used in biomedical research. An inbred strain is the one which has been maintained by sibling mating for twenty or more consecutive generations. Except for the sex difference, an inbred strain is homozygous at all loci. Most of the traits are consistent, do not vary in offspring and respond to experimental procedures uniformly. This enables researchers to produce reliable data for therapeutic purposes, hence inbred strain must be genetically pure over the generation. Methods: A total of 20 mice samples from 10 inbred strains i.e., A/J, BALB/c, C3H/J, CD1, C57BL/6, DBA/2, FVB/NJ, ICRC, Swiss Webster and Swiss/Ba were screened for their genetic purity with the help microsatellite markers. Tail DNA was isolated using Proteinase/K-Phenol-chloroform extraction method and quantified by Nanodrop. DNA obtained was used for microsatellite marker analysis by standardizing multiplex-PCR. Seven multiplex (duplex) panels were successfully established by optimizing various reaction conditions to analyse any deviation in the genetic profile. Result: Upon comparing genetic profiles obtained in multiplex PCR with the information available on Mouse Genome Informatics database, it was concluded that most of the inbred mice are genetically consistent as they showed zero divergence in observed base pair size mainly due to stringent breeding protocol and optimum living conditions. The work done also signifies a cost-effective, time saving, high throughput and robust method of genetic monitoring of laboratory mice strains.

Effects of Human RelA Transgene on Murine Macrophage Inflammatory Responses.

The NFκB transcription factors are major regulators of innate immune responses, and NFκB signal pathway dysregulation is linked to inflammatory disease. Here, we utilised bone marrow-derived macrophages from the p65-DsRedxp/IκBα-eGFP transgenic strain to study the functional implication of xenogeneic (human) RelA(p65) protein introduced into the mouse genome. Confocal imaging showed that human RelA is expressed in the cells and can translocate to the nucleus following activation of Toll-like receptor 4. RNA sequencing of lipid A-stimulated macrophages, revealed that human RelA impacts on murine gene transcription, affecting both non-NFκB and NFκB target genes, including immediate-early and late response genes, e.g., Fos and Cxcl10. Validation experiments on NFκB targets revealed markedly reduced mRNA levels, but similar kinetic profiles in transgenic cells compared to wild-type. Enrichment pathway analysis of differentially expressed genes revealed interferon and cytokine signaling were affected. These immune response pathways were also affected in macrophages treated with tumor necrosis factor. Data suggests that the presence of xenogeneic RelA protein likely has inhibitory activity, altering specific transcriptional profiles of key molecules involved in immune responses. It is therefore essential that this information be taken into consideration when designing and interpreting future experiments using this transgenic strain.

Copy Number Variation Analysis of Euploid Pregnancy Loss.

Objectives: Copy number variant (CNV) is believed to be the potential genetic cause of pregnancy loss. However, CNVs less than 3 Mb in euploid products of conceptions (POCs) remain largely unexplored. The aim of this study was to investigate the features of CNVs less than 3 Mb in POCs and their potential clinical significance in pregnancy loss/fetal death. Methods: CNV data were extracted from a cohort in our institution and 19 peer-reviewed publications, and only those CNVs less than 3 Mb detected in euploid pregnancy loss/fetal death were included. We conducted a CNV map to analyze the distribution of CNVs in chromosomes using R packages karyoploteR_1.10.5. Gene names and annotated gene types covered by those CNVs were mined from the human Release 19 reference genome file and GENECODE database. We assessed the expression patterns and the consequences of murine knock-out of those genes using TiGER and Mouse Genome Informatics (MGI) databases. Functional enrichment and pathway analysis for genes in CNVs were performed using clusterProfiler V3.12.0. Result: Breakpoints of 564 CNVs less than 3 Mb were obtained from 442 euploid POCs, with 349 gains and 185 losses. The CNV map showed that CNVs were distributed in all chromosomes, with the highest frequency detected in chromosome 22 and the lowest frequency in chromosome Y, and CNVs showed a higher density in the pericentromeric and sub-telomeric regions. A total of 5,414 genes mined from the CNV regions (CNVRs), Gene Ontology (GO), and pathway analysis showed that the genes were significantly enriched in multiple terms, especially in sensory perception, membrane region, and tight junction. A total of 995 protein-coding genes have been reported to present mammalian phenotypes in MGI, and 276 of them lead to embryonic lethality or abnormal embryo/placenta in knock-out mouse models. CNV located at 19p13.3 was the most common CNV of all POCs. Conclusion: CNVs less than 3 Mb in euploid POCs distribute unevenly in all chromosomes, and a higher density was seen in the pericentromeric and sub-telomeric regions. The genes in those CNVRs are significantly enriched in biological processes and pathways that are important to embryonic/fetal development. CNV in 19p13.3 and the variations of ARID3A and FSTL3 might contribute to pregnancy loss.

Hypothalamic control of energy expenditure and thermogenesis.

Energy expenditure and energy intake need to be balanced to maintain proper energy homeostasis. Energy homeostasis is tightly regulated by the central nervous system, and the hypothalamus is the primary center for the regulation of energy balance. The hypothalamus exerts its effect through both humoral and neuronal mechanisms, and each hypothalamic area has a distinct role in the regulation of energy expenditure. Recent studies have advanced the understanding of the molecular regulation of energy expenditure and thermogenesis in the hypothalamus with targeted manipulation techniques of the mouse genome and neuronal function. In this review, we elucidate recent progress in understanding the mechanism of how the hypothalamus affects basal metabolism, modulates physical activity, and adapts to environmental temperature and food intake changes.

DNAcycP: a deep learning tool for DNA cyclizability prediction.

DNA mechanical properties play a critical role in every aspect of DNA-dependent biological processes. Recently a high throughput assay named loop-seq has been developed to quantify the intrinsic bendability of a massive number of DNA fragments simultaneously. Using the loop-seq data, we develop a software tool, DNAcycP, based on a deep-learning approach for intrinsic DNA cyclizability prediction. We demonstrate DNAcycP predicts intrinsic DNA cyclizability with high fidelity compared to the experimental data. Using an independent dataset from in vitro selection for enrichment of loopable sequences, we further verified the predicted cyclizability score, termed C-score, can well distinguish DNA fragments with different loopability. We applied DNAcycP to multiple species and compared the C-scores with available high-resolution chemical nucleosome maps. Our analyses showed that both yeast and mouse genomes share a conserved feature of high DNA bendability spanning nucleosome dyads. Additionally, we extended our analysis to transcription factor binding sites and surprisingly found that the cyclizability is substantially elevated at CTCF binding sites in the mouse genome. We further demonstrate this distinct mechanical property is conserved across mammalian species and is inherent to CTCF binding DNA motif.

Discordance in STING-Induced Activation and Cell Death Between Mouse and Human Dendritic Cell Populations.

Stimulator of Interferon Genes (STING) is a cytosolic sensor of cyclic dinucleotides (CDNs). The activation of dendritic cells (DC) via the STING pathway, and their subsequent production of type I interferon (IFN) is considered central to eradicating tumours in mouse models. However, this contribution of STING in preclinical murine studies has not translated into positive outcomes of STING agonists in phase I & II clinical trials. We therefore questioned whether a difference in human DC responses could be critical to the lack of STING agonist efficacy in human settings. This study sought to directly compare mouse and human plasmacytoid DCs and conventional DC subset responses upon STING activation. We found all mouse and human DC subsets were potently activated by STING stimulation. As expected, Type I IFNs were produced by both mouse and human plasmacytoid DCs. However, mouse and human plasmacytoid and conventional DCs all produced type III IFNs (i.e., IFN-λs) in response to STING activation. Of particular interest, all human DCs produced large amounts of IFN-λ1, not expressed in the mouse genome. Furthermore, we also found differential cell death responses upon STING activation, observing rapid ablation of mouse, but not human, plasmacytoid DCs. STING-induced cell death in murine plasmacytoid DCs occurred in a cell-intrinsic manner and involved intrinsic apoptosis. These data highlight discordance between STING IFN and cell death responses in mouse and human DCs and caution against extrapolating STING-mediated events in mouse models to equivalent human outcomes.

Harmonizing model organism data in the Alliance of Genome Resources.

The Alliance of Genome Resources (the Alliance) is a combined effort of 7 knowledgebase projects: Saccharomyces Genome Database, WormBase, FlyBase, Mouse Genome Database, the Zebrafish Information Network, Rat Genome Database, and the Gene Ontology Resource. The Alliance seeks to provide several benefits: better service to the various communities served by these projects; a harmonized view of data for all biomedical researchers, bioinformaticians, clinicians, and students; and a more sustainable infrastructure. The Alliance has harmonized cross-organism data to provide useful comparative views of gene function, gene expression, and human disease relevance. The basis of the comparative views is shared calls of orthology relationships and the use of common ontologies. The key types of data are alleles and variants, gene function based on gene ontology annotations, phenotypes, association to human disease, gene expression, protein–protein and genetic interactions, and participation in pathways. The information is presented on uniform gene pages that allow facile summarization of information about each gene in each of the 7 organisms covered (budding yeast, roundworm Caenorhabditis elegans, fruit fly, house mouse, zebrafish, brown rat, and human). The harmonized knowledge is freely available on the alliancegenome.org portal, as downloadable files, and by APIs. We expect other existing and emerging knowledge bases to join in the effort to provide the union of useful data and features that each knowledge base currently provides.

Loss of chaperone-mediated autophagy is associated with low vertebral cancellous bone mass

Chaperone-mediated autophagy (CMA) is a protein degradation pathway that eliminates soluble cytoplasmic proteins that are damaged, incorrectly folded, or targeted for selective proteome remodeling. However, the role of CMA in skeletal homeostasis under physiological and pathophysiological conditions is unknown. To address the role of CMA for skeletal homeostasis, we deleted an essential component of the CMA process, namely Lamp2a, from the mouse genome. CRISPR-Cas9-based genome editing led to the deletion of both Lamp2a and Lamp2c, another Lamp2 isoform, producing Lamp2AC global knockout (L2ACgKO) mice. At 5 weeks of age female L2ACgKO mice had lower vertebral cancellous bone mass compared to wild-type (WT) controls, whereas there was no difference between genotypes in male mice at this age. The low bone mass of L2ACgKO mice was associated with elevated RANKL expression and the osteoclast marker genes Trap and Cathepsin K. At 18 weeks of age, both male and female L2ACgKO mice had lower vertebral cancellous bone mass compared to WT controls. The low bone mass of L2ACgKO mice was associated with increased osteoclastogenesis and decreased mineral deposition in cultured cells. Consistent with these findings, specific knockdown of Lamp2a in an osteoblastic cell line increased RANKL expression and decreased mineral deposition. Moreover, similar to what has been observed in other cell types, macroautophagy and proteasomal degradation were upregulated in CMA-deficient osteoblasts in culture. Thus, an increase in other protein degradation pathways may partially compensate for the loss of CMA in osteoblasts. Taken together, our results suggest that CMA plays a role in vertebral cancellous bone mass accrual in young adult mice and that this may be due to an inhibitory role of CMA on osteoclastogenesis or a positive role of CMA in osteoblast formation or function.

Mouse Gastric Epithelial Cells Resist CagA Delivery by the Helicobacter pylori Type IV Secretion System.

The initial step in bacterial infection is adherence of the bacterium to the target cell surface. Helicobacter pylori exploits the interaction of bacterial adhesin protein HopQ with human epithelial CEACAMs (CEACAM1, 5, and 6) to stably adhere to gastric epithelial cells, which is necessary for delivery of the H. pylori CagA oncoprotein into the epithelial cells via a type IV secretion system. In contrast to human CEACAMs, however, HopQ does not interact with Ceacam1 (mouse CEACAM1) in vitro or in CHO cells ectopically expressing Ceacam1. Since the mouse genome lacks Ceacam5 and Ceacam6, no significant HopQ–Ceacam interaction may occur in mouse gastric epithelial cells. Here, we found that the mouse stomach has a much lower expression level of Ceacam1 than the expression level of CEACAM1 in the human stomach. Consistently, mouse gastric epithelial cells resist CagA delivery by cagA-positive H. pylori, and the delivery is restored by ectopic expression of human CEACAM1 or CEACAM5 in mouse gastric epithelial cells. Thus, despite the fact that mice are routinely used for H. pylori infection studies, a low expression level of Ceacam1 in the mouse stomach together with the loss or greatly reduced interaction of HopQ with Ceacams make the mouse an inappropriate model for studying the role of H. pylori-delivered CagA in gastric pathogenesis, including the development of gastric cancer.

The mouse resource at National Resource Center for Mutant Mice.

Mouse models are essential for dissecting disease mechanisms and defining potential drug targets. There are more than 18,500 mouse strains available for research communities in National Resource Center for Mutant Mice (NRCMM) of China, affiliated with Model Animal Research Center of Nanjing University and Gempharmatech Company. In 2019, Gempharmatech launched the Knockout All Project (KOAP) aiming to generate null mutants and gene floxed strains for all protein-coding genes in mouse genome within 5years. So far, KOAP has generated 8,004 floxed strains and 9,769 KO (knockout) strains (updated to Oct, 2021). NRCMM also created hundreds of Cre transgenic lines, mutant knock-in models, immuno-deficient models, and humanized mouse models. As a member of the international mouse phenotyping consortium (IMPC), NRCMM provides comprehensive phenotyping services for mouse models. In summary, NRCMM will continue to support biomedical community with new mouse models as well as related services.

Patient-Specific TBX5-G125R Variant Induces Profound Transcriptional Deregulation and Atrial Dysfunction.

The pathogenic missense variant p.G125R in TBX5 (T-box transcription factor 5) causes Holt-Oram syndrome (also known as hand-heart syndrome) and early onset of atrial fibrillation. Revealing how an altered key developmental transcription factor modulates cardiac physiology in vivo will provide unique insights into the mechanisms underlying atrial fibrillation in these patients. We analyzed ECGs of an extended family pedigree of Holt-Oram syndrome patients. Next, we introduced the TBX5-p.G125R variant in the mouse genome (Tbx5G125R) and performed electrophysiologic analyses (ECG, optical mapping, patch clamp, intracellular calcium measurements), transcriptomics (single-nuclei and tissue RNA sequencing), and epigenetic profiling (assay for transposase-accessible chromatin using sequencing, H3K27ac [histone H3 lysine 27 acetylation] CUT&RUN [cleavage under targets and release under nuclease sequencing]). We discovered high incidence of atrial extra systoles and atrioventricular conduction disturbances in Holt-Oram syndrome patients. Tbx5G125R/+ mice were morphologically unaffected and displayed variable RR intervals, atrial extra systoles, and susceptibility to atrial fibrillation, reminiscent of TBX5-p.G125R patients. Atrial conduction velocity was not affected but systolic and diastolic intracellular calcium concentrations were decreased and action potentials were prolonged in isolated cardiomyocytes of Tbx5G125R/+ mice compared with controls. Transcriptional profiling of atria revealed the most profound transcriptional changes in cardiomyocytes versus other cell types, and identified over a thousand coding and noncoding transcripts that were differentially expressed. Epigenetic profiling uncovered thousands of TBX5-p.G125R-sensitive, putative regulatory elements (including enhancers) that gained accessibility in atrial cardiomyocytes. The majority of sites with increased accessibility were occupied by Tbx5. The small group of sites with reduced accessibility was enriched for DNA-binding motifs of members of the SP (specificity protein) and KLF (Krüppel-like factor) families of transcription factors. These data show that Tbx5-p.G125R induces changes in regulatory element activity, alters transcriptional regulation, and changes cardiomyocyte behavior, possibly caused by altered DNA binding and cooperativity properties. Our data reveal that a disease-causing missense variant in TBX5 induces profound changes in the atrial transcriptional regulatory network and epigenetic state in vivo, leading to arrhythmia reminiscent of those seen in human TBX5-p.G125R variant carriers.

Host-pathogen genetic interactions underlie tuberculosis susceptibility in genetically diverse mice.

The outcome of an encounter with Mycobacterium tuberculosis (Mtb) depends on the pathogen's ability to adapt to the variable immune pressures exerted by the host. Understanding this interplay has proven difficult, largely because experimentally tractable animal models do not recapitulate the heterogeneity of tuberculosis disease. We leveraged the genetically diverse Collaborative Cross (CC) mouse panel in conjunction with a library of Mtb mutants to create a resource for associating bacterial genetic requirements with host genetics and immunity. We report that CC strains vary dramatically in their susceptibility to infection and produce qualitatively distinct immune states. Global analysis of Mtb transposon mutant fitness (TnSeq) across the CC panel revealed that many virulence pathways are only required in specific host microenvironments, identifying a large fraction of the pathogen's genome that has been maintained to ensure fitness in a diverse population. Both immunological and bacterial traits can be associated with genetic variants distributed across the mouse genome, making the CC a unique population for identifying specific host-pathogen genetic interactions that influence pathogenesis.

Functional and genomic comparative study of the bitter taste receptor family TAS2R: Insight into the role of human TAS2R5.

Bitterness is perceived in humans by 25subtypes of bitter taste receptors (hTAS2R) that range from broadly tuned to more narrowly tuned receptors. hTAS2R5 is one of the most narrowly tuned bitter taste receptors in humans. In this study, we review the literature on this receptor and show there is no consensus about its role. We then compare the possible role of hTAS2R5 with that of the proteins of the TAS2R family in rat, mouse, and pig. A phylogenetic tree of all mammalian TAS2R domain-containing proteins showed that human hTAS2R5has no ortholog in pig, mouse, or rat genomes. By comparing the agonists that are common to hTAS2R5 and other members of the family, we observed that hTAS2R39 is the receptor that shares most agonists with hTAS2R5. In mouse, some of these agonists activate mTas2r105 and mTas2r144, which are distant paralogs of hTAS2R5. mTas2r144seems to be the receptor that is most similar to hTAS2R5 because they are both activated by the same agonists and have affinities in the same range of values. Then, we can conclude that hTAS2R5has a unique functional specificity in humans as it is activated by selective agonists and that its closest functional homolog in mouse is the phylogenetically distant mTas2r144.

The dark matter of the human genome and its role in human cancers.

The transcribed ultra-conserved regions (T-UCRs) are a novel family of non-coding RNAs which are absolutely conserved (100%) across orthologous regions of the human, mouse, and rat genomes. T-UCRs represent a small portion of the human genome that is likely to be functional but does not code for proteins and is referred to as the "dark matter" of the human genome. Although T-UCRs are ubiquitously expressed, tissue- and disease-specific expression of T-UCRs have also been observed. Accumulating evidence suggests that T-UCRs are differentially expressed and involved in the malignant transformation of human tumors through various genetic and epigenetic regulatory mechanisms. Therefore, T-UCRs are novel candidate predisposing biomarkers for cancer development. T-UCRs have shown to drive malignant transformation of human cancers through regulating non-coding RNAs and/or protein coding genes. However, the functions and fate of most T-UCRs remain mysterious. Here, we review and highlight the current knowledge on these ultra-conserved elements in the formation and progression of human cancers.

MOET: a web-based gene set enrichment tool at the Rat Genome Database for multiontology and multispecies analyses.

Biological interpretation of a large amount of gene or protein data is complex. Ontology analysis tools are imperative in finding functional similarities through overrepresentation or enrichment of terms associated with the input gene or protein lists. However, most tools are limited by their ability to do ontology-specific and species-limited analyses. Furthermore, some enrichment tools are not updated frequently with recent information from databases, thus giving users inaccurate, outdated or uninformative data. Here, we present MOET or the Multi-Ontology Enrichment Tool (v.1 released in April 2019 and v.2 released in May 2021), an ontology analysis tool leveraging data that the Rat Genome Database (RGD) integrated from in-house expert curation and external databases including the National Center for Biotechnology Information (NCBI), Mouse Genome Informatics (MGI), The Kyoto Encyclopedia of Genes and Genomes (KEGG), The Gene Ontology Resource, UniProt-GOA, and others. Given a gene or protein list, MOET analysis identifies significantly overrepresented ontology terms using a hypergeometric test and provides nominal and Bonferroni corrected P-values and odds ratios for the overrepresented terms. The results are shown as a downloadable list of terms with and without Bonferroni correction, and a graph of the P-values and number of annotated genes for each term in the list. MOET can be accessed freely from https://rgd.mcw.edu/rgdweb/enrichment/start.html.