On-the-fly selection of cell-specific enhancers, genes, miRNAs and proteins across the human body using SlideBase

Genomics consortia have produced large datasets profiling the expression of genes, micro-RNAs, enhancers and more across human tissues or cells. There is a need for intuitive tools to select subsets of such data that is the most relevant for specific studies. To this end, we present SlideBase, a web tool which offers a new way of selecting genes, promoters, enhancers and microRNAs that are preferentially expressed/used in a specified set of cells/tissues, based on the use of interactive sliders. With the help of sliders, SlideBase enables users to define custom expression thresholds for individual cell types/tissues, producing sets of genes, enhancers etc. which satisfy these constraints. Changes in slider settings result in simultaneous changes in the selected sets, updated in real time. SlideBase is linked to major databases from genomics consortia, including FANTOM, GTEx, The Human Protein Atlas and BioGPS. Database URL: http://slidebase.binf.ku.d

IMOTA: an interactive multi-omics tissue atlas for the analysis of human miRNA-target interactions

Web repositories for almost all ‘omics’ types have been generated—detailing the repertoire of representatives across different tissues or cell types. A logical next step is the combination of these valuable sources. With IMOTA (interactive multi omics tissue atlas), we developed a database that includes 23 725 relations between miRNAs and 23 tissues, 310 932 relations between mRNAs and the same tissues as well as 63 043 relations between proteins and the 23 tissues in Homo sapiens. IMOTA also contains data on tissue-specific interactions, e.g. information on 331 413 miRNAs and target gene pairs that are jointly expressed in the considered tissues. By using intuitive filter and visualization techniques, it is with minimal effort possible to answer various questions. These include rather general questions but also requests specific for genes, miRNAs or proteins. An example for a general task could be ‘identify all miRNAs, genes and proteins in the lung that are highly expressed and where experimental evidence proves that the miRNAs target the genes’. An example for a specific request for a gene and a miRNA could for example be ‘In which tissues is miR-34c and its target gene BCL2 expressed?’. The IMOTA repository is freely available online at https://ccb-web.cs.uni-saarland.de/imota/

The intersectional genetics landscape for humans

BACKGROUND: The human body is made up of hundreds-perhaps thousands-of cell types and states, most of which are currently inaccessible genetically. Intersectional genetic approaches can increase the number of genetically accessible cells, but the scope and safety of these approaches have not been systematically assessed. A typical intersectional method acts like an "AND" logic gate by converting the input of 2 or more active, yet unspecific, regulatory elements (REs) into a single cell type specific synthetic output. RESULTS: Here, we systematically assessed the intersectional genetics landscape of the human genome using a subset of cells from a large RE usage atlas (Functional ANnoTation Of the Mammalian genome 5 consortium, FANTOM5) obtained by cap analysis of gene expression sequencing (CAGE-seq). We developed the heuristics and algorithms to retrieve and quality-rank "AND" gate intersections. Of the 154 primary cell types surveyed, >90% can be distinguished from each other with as few as 3 to 4 active REs, with quantifiable safety and robustness. We call these minimal intersections of active REs with cell-type diagnostic potential "versatile entry codes" (VEnCodes). Each of the 158 cancer cell types surveyed could also be distinguished from the healthy primary cell types with small VEnCodes, most of which were robust to intra- and interindividual variation. Methods for the cross-validation of CAGE-seq-derived VEnCodes and for the extraction of VEnCodes from pooled single-cell sequencing data are also presented. CONCLUSIONS: Our work provides a systematic view of the intersectional genetics landscape in humans and demonstrates the potential of these approaches for future gene delivery technologies.publishersversionpublishe

Investigation into the genetic basis of bovine horn development

The presence of horns in ruminants has financial and welfare implications for the farming of cattle, sheep and goats worldwide. The genetic interactions that lead to horn development are not known. Hornless, or polled, cattle occur naturally, but the known causative DNA variants (Celtic, Friesian, Mongolian and Guarani) are in intergenic regions on bovine chromosome 1, and therefore, their functions are not known. The leading hypothesis is that horns are derived from cranial neural crest cells and the POLLED variants disrupt the migration or proliferation of these stem cells. The bovine POLLED region was explored through bioinformatics analyses as horned animals may have genomic differences from hornless individuals or species near the POLLED DNA variants. The aim was to identify differences in genes synteny, lincRNA, and topologically associating domain (TAD) structure between horned and hornless individuals or species. Horned (n = 1) and polled (Celtic; n = 1) Hi-C sequences produced the same TAD structures. The POLLED genomic region was refined to a 520-kb region encompassing all four POLLED variants. LOC526226 was unique to the bovine POLLED region and not conserved in the species analysed (water buffalo, sheep, goat, pig, horse, dog and human), and therefore, may be involved in horn development. Histological analyses of cranial tissues from homozygous horned and polled fetuses at day 58 of development were conducted. The aims were to 1) determine the differences in the structure of horn bud region, and 2) compare immunohistochemistry staining of neural crest markers (SOX10 and NGFR) and RXFP2 between horned and polled tissues. Condensed cells were only observed in the horn bud mesenchyme of horned fetuses and may be progenitor cells. SOX10 and NGFR was not detected in these condensed cells, and therefore, these cells are not derived from the neural crest or have differentiated and no longer express neural crest markers. SOX10 and NGFR were detected in the peripheral nerves. RXFP2 was detected in peripheral nerves and in the horn bud epidermis. Transcriptomic analyses of cranial tissues from the horned and polled fetuses at day 58 of development was also conducted. The aims were to 1) identify genes that may directly be affected by the polled variants, and 2) identify genes and pathways important for horn development. Near the POLLED region, three genes (C1H21orf62, SON and EVA1C) and one lincRNA (LOC112447120) were differentially expressed between horned and polled fetuses. Previously identified candidate genes, RXFP2, TWIST2 and ZEB2, were also differentially expressed. New candidates for the horn development pathway were proposed based on the analyses (MEIS2, PBX3, FZD8, CTNNB1 and LEF1). LOC526226 was not differentially expressed in the horn bud. Differentially expressed genes had functions in axon guidance, cytoskeletal structure and the extracellular region, and therefore, these pathways may be vital for horn development. Based on this research, it is now hypothesised that 1) horn stem cells are located in the mesenchyme and interact with the epidermis to initiate horn development, 2) the Celtic POLLED variant directly affects expression of C1H21orf62, SON, EVA1C and LOC112447120, and 3) the migration of horn stem cells is reduced by the effect of the POLLED variants upon C1H21orf62, SON, EVA1C and/or LOC112447120 expression.Thesis (Ph.D.) -- University of Adelaide, School of Animal and Veterinary Sciences, 202

VenCode – a versatile entry code for post-DNA delivery identification of target cells

RESUMO: O corpo humano é feito de centenas, talvez milhares de tipos e estados celulares distintos, a maioria atualmente inacessíveis através de ferramentas genéticas. A acessibilidade genética traz consigo um potencial terapêutico e de diagnostico significante, ao permitir a entrega seletiva de mensagens genéticas, ou terapias, diretamente às células. Trabalhos em organismos modelo mostram que a atividade de um só elemento regulatório (ER) é raramente específica para um tipo celular, o que limita o seu uso a sistemas genéticos desenhados para controlar a expressão genica após a sua entrega nas células. Abordagens de genética interseccional podem, em teoria, aumentar o número de células acessíveis sem esta restrição, mas o âmbito e a segurança dessas abordagens para o organismo humano não foram ainda sistematicamente estudados devido a uma falta de bases de dados de ERs para um extenso número de tipos celulares, e de métodos para as explorar. Um típico método interseccional funciona como uma porta logica AND ao converter a informação de dois ou mais ERs ativos num só sinal de saída, que se torna único para a célula analisada. Neste trabalho, estudamos sistematicamente o panorama da genética intersecional no organismo humano, usando um grupo de células selecionado a partir de um atlas de atividade de ERs obtido através do sequenciamento “Cap analysis of Gene Expression” (CAGE-seq) de centenas de células primárias e de cancro (o atlas do consórcio FANTOM5). Desenvolvemos algoritmos e heurísticas para encontrar e recolher intersecções do tipo porta AND e em seguida para determinar a sua qualidade. Descobrimos que mais de 90% dos 154 tipos celulares primários estudados podem ser distinguidos uns dos outros com apenas 3 ou 4 ERs ativos, com segurança e robustez. Chamamos de “Versatile Entry Codes” (VEnCodes) a esse número mínimo de intersecções de ERs ativos com potencial de distinção celular. Cada uma das 158 células cancerígenas estudadas poderam ser distinguidas do grupo de células saudáveis com VEnCodes de poucos ERs, a maioria dos quais são altamente robustos à variação intra- e inter-individual. Finalmente, fornecemos métodos para a validação dos VEnCodes obtidos e para a obtenção de VEnCodes a partir de bases de dados de sequenciamento ao nível de célula-a-célula. O nosso trabalho oferece uma visão sistemática do panorama da genética interseccional no organismo humano e demonstra o potencial dessas abordagens para tecnologias futuras de terapia genética.ABSTRACT: The human body is made up of hundreds, perhaps thousands of cell types and states, most of which are currently inaccessible genetically. Genetic accessibility carries significant diagnostic and therapeutic potential by allowing the selective delivery of genetic messages or cures to cells. Research in model organisms has shown that single regulatory element (RE) activities are seldom cell type specific, limiting their usage in genetic systems designed to restrict gene expression posteriorly to their delivery to cells. Intersectional genetic approaches can theoretically increase the number of genetically accessible cells, but the scope and safety of these approaches to human have not been systematically assessed due primarily to the lack of suitable thorough RE activity databases and methods to explore them. A typical intersectional method acts like an AND logic gate by converting the input of two or more active REs into a single synthetic output, which becomes unique for that cell. Here, we systematically assessed the intersectional genetics landscape of human using a curated subset of cells from a large RE usage atlas obtained by Cap Analysis of Gene Expression sequencing (CAGE-seq) of thousands of primary and cancer cells (the FANTOM5 consortium atlas). We developed the heuristics and algorithms to retrieve AND gate intersections and quality-rank them intra- and interindividually. We find that >90% of the 154 primary cell types surveyed can be distinguished from each other with as little as 3 to 4 active REs, with quantifiable safety and robustness. We call these minimal intersections of active REs with cell-type diagnostic potential "Versatile Entry Codes" (VEnCodes). Each of the 158 cancer cell types surveyed could also be distinguished from the healthy primary cell types with small VEnCodes, most of which were highly robust to intra- and interindividual variation. Finally, we provide methods for the cross-validation of CAGE-seq-derived VEnCodes and for the extraction of VEnCodes from pooled single cell sequencing data. Our work provides a systematic view of the intersectional genetics landscape in human and demonstrates the potential of these approaches for future gene delivery technologies in human