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Article ; Online: Computational Integration of HSV-1 Multi-omics Data.

Friedel, Caroline C

Methods in molecular biology (Clifton, N.J.)

2022 Volume 2610, Page(s) 31–48

Abstract: Functional genomics techniques based on next-generation sequencing provide new avenues for studying host responses to viral infections at multiple levels, including transcriptional and translational processes and chromatin organization. This chapter ... ...

Abstract	Functional genomics techniques based on next-generation sequencing provide new avenues for studying host responses to viral infections at multiple levels, including transcriptional and translational processes and chromatin organization. This chapter provides an overview on the computational integration of multiple types of "omics" data on lytic herpes simplex virus 1 (HSV-1) infection. It summarizes methods developed and applied in two publications that combined 4sU-seq for studying de novo transcription, ribosome profiling for investigating active translation, RNA-seq of subcellular RNA fractions for determining subcellular location of transcripts, and ATAC-seq for profiling chromatin accessibility genome-wide. These studies revealed an unprecedented disruption of transcription termination in HSV-1 infection resulting in widespread read-through transcription beyond poly(A) sites for most but not all host genes. This impacts chromatin architecture by increasing chromatin accessibility selectively in downstream regions of affected genes. In this way, computational integration of multi-omics data identified novel and unsuspected mechanisms at play in lytic HSV-1 infection.
MeSH term(s)	Humans ; Chromatin ; Herpes Simplex ; Herpesvirus 1, Human/genetics ; Multiomics ; Transcription, Genetic
Chemical Substances	Chromatin
Language	English
Publishing date	2022-12-19
Publishing country	United States
Document type	Journal Article ; Review
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-0716-2895-9_3
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Article ; Online: RegCFinder: targeted discovery of genomic subregions with differential read density.

Weiß, Elena / Friedel, Caroline C

Bioinformatics advances

2023 Volume 3, Issue 1, Page(s) vbad085

Abstract: Motivation: To date, no methods are available for the targeted identification of genomic subregions with differences in sequencing read distributions between two conditions. Existing approaches either only determine absolute read number changes, require ...

Abstract	Motivation: To date, no methods are available for the targeted identification of genomic subregions with differences in sequencing read distributions between two conditions. Existing approaches either only determine absolute read number changes, require predefined subdivisions of input windows or average across multiple genes. Results: Here, we present RegCFinder, which automatically identifies subregions of input windows with differences in read density between two conditions. For this purpose, the problem is defined as an instance of the all maximum scoring subsequences problem, which can be solved in linear time. Subsequently, statistical significance and differential usage of identified subregions are determined with DEXSeq. RegCFinder allows flexible definition of input windows to target the analysis to any regions of interests, e.g. promoters, gene bodies, peak regions and more. Furthermore, any type of sequencing assay can be used as input; thus, RegCFinder lends itself to a wide range of applications. We illustrate the usefulness of RegCFinder on two applications, where we can both confirm previous results and identify interesting gene subgroups with distinctive changes in read distributions. Availability and implementation: RegCFinder is implemented as a workflow for the workflow management system Watchdog and available at: https://github.com/watchdog-wms/watchdog-wms-workflows/. Supplementary information: Supplementary data are available at
Language	English
Publishing date	2023-07-04
Publishing country	England
Document type	Journal Article
ISSN	2635-0041
ISSN (online)	2635-0041
DOI	10.1093/bioadv/vbad085
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Article ; Online: Inhibition of CDK12 elevates cancer cell dependence on P-TEFb by stimulation of RNA polymerase II pause release.

Wang, Zhijia / Himanen, Samu V / Haikala, Heidi M / Friedel, Caroline C / Vihervaara, Anniina / Barborič, Matjaž

Nucleic acids research

2023 Volume 51, Issue 20, Page(s) 10970–10991

Abstract: P-TEFb and CDK12 facilitate transcriptional elongation by RNA polymerase II. Given the prominence of both kinases in cancer, gaining a better understanding of their interplay could inform the design of novel anti-cancer strategies. While down-regulation ... ...

Abstract	P-TEFb and CDK12 facilitate transcriptional elongation by RNA polymerase II. Given the prominence of both kinases in cancer, gaining a better understanding of their interplay could inform the design of novel anti-cancer strategies. While down-regulation of DNA repair genes in CDK12-targeted cancer cells is being explored therapeutically, little is known about mechanisms and significance of transcriptional induction upon inhibition of CDK12. We show that selective targeting of CDK12 in colon cancer-derived cells activates P-TEFb via its release from the inhibitory 7SK snRNP. In turn, P-TEFb stimulates Pol II pause release at thousands of genes, most of which become newly dependent on P-TEFb. Amongst the induced genes are those stimulated by hallmark pathways in cancer, including p53 and NF-κB. Consequently, CDK12-inhibited cancer cells exhibit hypersensitivity to inhibitors of P-TEFb. While blocking P-TEFb triggers their apoptosis in a p53-dependent manner, it impedes cell proliferation irrespective of p53 by preventing induction of genes downstream of the DNA damage-induced NF-κB signaling. In summary, stimulation of Pol II pause release at the signal-responsive genes underlies the functional dependence of CDK12-inhibited cancer cells on P-TEFb. Our study establishes the mechanistic underpinning for combinatorial targeting of CDK12 with either P-TEFb or the induced oncogenic pathways in cancer.
MeSH term(s)	Neoplasms/genetics ; NF-kappa B/genetics ; NF-kappa B/metabolism ; Positive Transcriptional Elongation Factor B/genetics ; Positive Transcriptional Elongation Factor B/metabolism ; Ribonucleoproteins, Small Nuclear/genetics ; RNA Polymerase II/genetics ; RNA Polymerase II/metabolism ; RNA-Binding Proteins/metabolism ; Tumor Suppressor Protein p53/genetics ; Humans ; Cell Line, Tumor
Chemical Substances	NF-kappa B ; Positive Transcriptional Elongation Factor B (EC 2.7.11.-) ; Ribonucleoproteins, Small Nuclear ; RNA Polymerase II (EC 2.7.7.-) ; RNA-Binding Proteins ; Tumor Suppressor Protein p53
Language	English
Publishing date	2023-10-09
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkad792
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Article ; Online: Watchdog - a workflow management system for the distributed analysis of large-scale experimental data.

Kluge, Michael / Friedel, Caroline C

BMC bioinformatics

2018 Volume 19, Issue 1, Page(s) 97

Abstract: Background: The development of high-throughput experimental technologies, such as next-generation sequencing, have led to new challenges for handling, analyzing and integrating the resulting large and diverse datasets. Bioinformatical analysis of these ... ...

Abstract	Background: The development of high-throughput experimental technologies, such as next-generation sequencing, have led to new challenges for handling, analyzing and integrating the resulting large and diverse datasets. Bioinformatical analysis of these data commonly requires a number of mutually dependent steps applied to numerous samples for multiple conditions and replicates. To support these analyses, a number of workflow management systems (WMSs) have been developed to allow automated execution of corresponding analysis workflows. Major advantages of WMSs are the easy reproducibility of results as well as the reusability of workflows or their components. Results: In this article, we present Watchdog, a WMS for the automated analysis of large-scale experimental data. Main features include straightforward processing of replicate data, support for distributed computer systems, customizable error detection and manual intervention into workflow execution. Watchdog is implemented in Java and thus platform-independent and allows easy sharing of workflows and corresponding program modules. It provides a graphical user interface (GUI) for workflow construction using pre-defined modules as well as a helper script for creating new module definitions. Execution of workflows is possible using either the GUI or a command-line interface and a web-interface is provided for monitoring the execution status and intervening in case of errors. To illustrate its potentials on a real-life example, a comprehensive workflow and modules for the analysis of RNA-seq experiments were implemented and are provided with the software in addition to simple test examples. Conclusions: Watchdog is a powerful and flexible WMS for the analysis of large-scale high-throughput experiments. We believe it will greatly benefit both users with and without programming skills who want to develop and apply bioinformatical workflows with reasonable overhead. The software, example workflows and a comprehensive documentation are freely available at www.bio.ifi.lmu.de/watchdog.
MeSH term(s)	Computational Biology/methods ; Herpes Simplex/genetics ; Herpes Simplex/virology ; Herpesvirus 1, Human/genetics ; High-Throughput Nucleotide Sequencing/methods ; Humans ; RNA/analysis ; RNA/genetics ; Software ; User-Computer Interface ; Virus Replication ; Workflow
Chemical Substances	RNA (63231-63-0)
Language	English
Publishing date	2018-03-13
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2041484-5
ISSN	1471-2105 ; 1471-2105
ISSN (online)	1471-2105
ISSN	1471-2105
DOI	10.1186/s12859-018-2107-4
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Article ; Online: Computational analysis of virus-host interactomes.

Friedel, Caroline C

Methods in molecular biology (Clifton, N.J.)

2013 Volume 1064, Page(s) 115–130

Abstract: High-throughput methods for screening of physical and functional interactions now provide the means to study virus-host interactions on a genome scale. The limited coverage of these methods and the large size and uncertain quality of the identified ... ...

Abstract	High-throughput methods for screening of physical and functional interactions now provide the means to study virus-host interactions on a genome scale. The limited coverage of these methods and the large size and uncertain quality of the identified interaction sets, however, require sophisticated computational approaches to obtain novel insights and hypotheses on virus infection processes from these interactions. Here, we describe the central steps of bioinformatics methods applied most commonly for this task and highlight important aspects that need to be considered and potential pitfalls that should be avoided.
MeSH term(s)	Animals ; Computational Biology/methods ; Databases, Genetic ; Host-Pathogen Interactions ; Humans ; Infections/metabolism ; Infections/virology ; Protein Interaction Mapping/methods ; Proteomics/methods ; Two-Hybrid System Techniques ; Virus Diseases/metabolism
Keywords	covid19
Language	English
Publishing date	2013-07-23
Publishing country	United States
Document type	Journal Article
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-62703-601-6_8
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Article ; Online: Prediction of Poly(A) Sites by Poly(A) Read Mapping.

Bonfert, Thomas / Friedel, Caroline C

PloS one

2017 Volume 12, Issue 1, Page(s) e0170914

Abstract: RNA-seq reads containing part of the poly(A) tail of transcripts (denoted as poly(A) reads) provide the most direct evidence for the position of poly(A) sites in the genome. However, due to reduced coverage of poly(A) tails by reads, poly(A) reads are ... ...

Abstract	RNA-seq reads containing part of the poly(A) tail of transcripts (denoted as poly(A) reads) provide the most direct evidence for the position of poly(A) sites in the genome. However, due to reduced coverage of poly(A) tails by reads, poly(A) reads are not routinely identified during RNA-seq mapping. Nevertheless, recent studies for several herpesviruses successfully employed mapping of poly(A) reads to identify herpesvirus poly(A) sites using different strategies and customized programs. To more easily allow such analyses without requiring additional programs, we integrated poly(A) read mapping and prediction of poly(A) sites into our RNA-seq mapping program ContextMap 2. The implemented approach essentially generalizes previously used poly(A) read mapping approaches and combines them with the context-based approach of ContextMap 2 to take into account information provided by other reads aligned to the same location. Poly(A) read mapping using ContextMap 2 was evaluated on real-life data from the ENCODE project and compared against a competing approach based on transcriptome assembly (KLEAT). This showed high positive predictive value for our approach, evidenced also by the presence of poly(A) signals, and considerably lower runtime than KLEAT. Although sensitivity is low for both methods, we show that this is in part due to a high extent of spurious results in the gold standard set derived from RNA-PET data. Sensitivity improves for poly(A) sites of known transcripts or determined with a more specific poly(A) sequencing protocol and increases with read coverage on transcript ends. Finally, we illustrate the usefulness of the approach in a high read coverage scenario by a re-analysis of published data for herpes simplex virus 1. Thus, with current trends towards increasing sequencing depth and read length, poly(A) read mapping will prove to be increasingly useful and can now be performed automatically during RNA-seq mapping with ContextMap 2.
MeSH term(s)	Animals ; Humans ; Poly A/genetics ; RNA, Messenger/genetics ; Sequence Analysis, RNA/methods ; Software ; Transcriptome/genetics
Chemical Substances	RNA, Messenger ; Poly A (24937-83-5)
Language	English
Publishing date	2017
Publishing country	United States
Document type	Journal Article
ISSN	1932-6203
ISSN (online)	1932-6203
DOI	10.1371/journal.pone.0170914
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Article ; Online: P-TEFb promotes cell survival upon p53 activation by suppressing intrinsic apoptosis pathway.

Wang, Zhijia / Mačáková, Monika / Bugai, Andrii / Kuznetsov, Sergey G / Hassinen, Antti / Lenasi, Tina / Potdar, Swapnil / Friedel, Caroline C / Barborič, Matjaž

Nucleic acids research

2023 Volume 51, Issue 4, Page(s) 1687–1706

Abstract: Positive transcription elongation factor b (P-TEFb) is the crucial player in RNA polymerase II (Pol II) pause release that has emerged as a promising target in cancer. Because single-agent therapy may fail to deliver durable clinical response, targeting ... ...

Abstract	Positive transcription elongation factor b (P-TEFb) is the crucial player in RNA polymerase II (Pol II) pause release that has emerged as a promising target in cancer. Because single-agent therapy may fail to deliver durable clinical response, targeting of P-TEFb shall benefit when deployed as a combination therapy. We screened a comprehensive oncology library and identified clinically relevant antimetabolites and Mouse double minute 2 homolog (MDM2) inhibitors as top compounds eliciting p53-dependent death of colorectal cancer cells in synergy with selective inhibitors of P-TEFb. While the targeting of P-TEFb augments apoptosis by anti-metabolite 5-fluorouracil, it switches the fate of cancer cells by the non-genotoxic MDM2 inhibitor Nutlin-3a from cell-cycle arrest to apoptosis. Mechanistically, the fate switching is enabled by the induction of p53-dependent pro-apoptotic genes and repression of P-TEFb-dependent pro-survival genes of the PI3K-AKT signaling cascade, which stimulates caspase 9 and intrinsic apoptosis pathway in BAX/BAK-dependent manner. Finally, combination treatments trigger apoptosis of cancer cell spheroids. Together, co-targeting of P-TEFb and suppressors of intrinsic apoptosis could become a viable strategy to eliminate cancer cells.
MeSH term(s)	Apoptosis ; Cell Line, Tumor ; Cell Survival ; Phosphatidylinositol 3-Kinases/metabolism ; Positive Transcriptional Elongation Factor B/antagonists & inhibitors ; Positive Transcriptional Elongation Factor B/metabolism ; Proto-Oncogene Proteins c-mdm2/genetics ; Tumor Suppressor Protein p53/genetics ; Humans
Chemical Substances	Phosphatidylinositol 3-Kinases (EC 2.7.1.-) ; Positive Transcriptional Elongation Factor B (EC 2.7.11.-) ; Proto-Oncogene Proteins c-mdm2 (EC 2.3.2.27) ; Tumor Suppressor Protein p53 ; MDM2 protein, human (EC 2.3.2.27)
Language	English
Publishing date	2023-02-01
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkad001
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Article ; Online: Watchdog 2.0: New developments for reusability, reproducibility, and workflow execution.

Kluge, Michael / Friedl, Marie-Sophie / Menzel, Amrei L / Friedel, Caroline C

GigaScience

2020 Volume 9, Issue 6

Abstract: Background: Advances in high-throughput methods have brought new challenges for biological data analysis, often requiring many interdependent steps applied to a large number of samples. To address this challenge, workflow management systems, such as ... ...

Abstract	Background: Advances in high-throughput methods have brought new challenges for biological data analysis, often requiring many interdependent steps applied to a large number of samples. To address this challenge, workflow management systems, such as Watchdog, have been developed to support scientists in the (semi-)automated execution of large analysis workflows. Implementation: Here, we present Watchdog 2.0, which implements new developments for module creation, reusability, and documentation and for reproducibility of analyses and workflow execution. Developments include a graphical user interface for semi-automatic module creation from software help pages, sharing repositories for modules and workflows, and a standardized module documentation format. The latter allows generation of a customized reference book of public and user-specific modules. Furthermore, extensive logging of workflow execution, module and software versions, and explicit support for package managers and container virtualization now ensures reproducibility of results. A step-by-step analysis protocol generated from the log file may, e.g., serve as a draft of a manuscript methods section. Finally, 2 new execution modes were implemented. One allows resuming workflow execution after interruption or modification without rerunning successfully executed tasks not affected by changes. The second one allows detaching and reattaching to workflow execution on a local computer while tasks continue running on computer clusters. Conclusions: Watchdog 2.0 provides several new developments that we believe to be of benefit for large-scale bioinformatics analysis and that are not completely covered by other competing workflow management systems. The software itself, module and workflow repositories, and comprehensive documentation are freely available at https://www.bio.ifi.lmu.de/watchdog.
MeSH term(s)	Algorithms ; Computational Biology/methods ; Computational Biology/standards ; High-Throughput Nucleotide Sequencing ; Reproducibility of Results ; Software ; User-Computer Interface ; Workflow
Language	English
Publishing date	2020-06-17
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2708999-X
ISSN	2047-217X ; 2047-217X
ISSN (online)	2047-217X
ISSN	2047-217X
DOI	10.1093/gigascience/giaa068
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Article ; Online: HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform.

Friedl, Marie-Sophie / Djakovic, Lara / Kluge, Michael / Hennig, Thomas / Whisnant, Adam W / Backes, Simone / Dölken, Lars / Friedel, Caroline C

PloS one

2022 Volume 17, Issue 10, Page(s) e0276467

Abstract: The herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs by a translation-initiation-dependent mechanism, which should spare circular RNAs (circRNAs). Here, we show that vhs-mediated degradation of ... ...

Abstract	The herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs by a translation-initiation-dependent mechanism, which should spare circular RNAs (circRNAs). Here, we show that vhs-mediated degradation of linear mRNAs leads to an enrichment of circRNAs relative to linear mRNAs during HSV-1 infection. This was also observed in influenza A virus (IAV) infection, likely due to degradation of linear host mRNAs mediated by the IAV PA-X protein and cap-snatching RNA-dependent RNA polymerase. For most circRNAs, enrichment was not due to increased circRNA synthesis but due to a general loss of linear RNAs. In contrast, biogenesis of a circRNA originating from the long isoform (NEAT1_2) of the nuclear paraspeckle assembly transcript 1 (NEAT1) was induced both in HSV-1 infection-in a vhs-independent manner-and in IAV infection. This was associated with induction of novel linear splicing of NEAT1_2 both within and downstream of the circRNA. NEAT1_2 forms a scaffold for paraspeckles, nuclear bodies located in the interchromatin space, must likely remain unspliced for paraspeckle assembly and is up-regulated in HSV-1 and IAV infection. We show that NEAT1_2 splicing and up-regulation can be induced by ectopic co-expression of the HSV-1 immediate-early proteins ICP22 and ICP27, potentially linking increased expression and splicing of NEAT1_2. To identify other conditions with NEAT1_2 splicing, we performed a large-scale screen of published RNA-seq data. This uncovered both induction of NEAT1_2 splicing and poly(A) read-through similar to HSV-1 and IAV infection in cancer cells upon inhibition or knockdown of CDK7 or the MED1 subunit of the Mediator complex phosphorylated by CDK7. In summary, our study reveals induction of novel circular and linear NEAT1_2 splicing isoforms as a common characteristic of HSV-1 and IAV infection and highlights a potential role of CDK7 in HSV-1 or IAV infection.
MeSH term(s)	Humans ; Herpesvirus 1, Human/genetics ; RNA, Circular ; Immediate-Early Proteins/genetics ; Influenza, Human ; Herpes Simplex ; RNA, Messenger/genetics ; Protein Isoforms/genetics ; RNA-Dependent RNA Polymerase ; Mediator Complex
Chemical Substances	RNA, Circular ; Immediate-Early Proteins ; RNA, Messenger ; Protein Isoforms ; RNA-Dependent RNA Polymerase (EC 2.7.7.48) ; Mediator Complex
Language	English
Publishing date	2022-10-24
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2267670-3
ISSN	1932-6203 ; 1932-6203
ISSN (online)	1932-6203
ISSN	1932-6203
DOI	10.1371/journal.pone.0276467
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Article ; Online: Decoding murine cytomegalovirus.

Lodha, Manivel / Muchsin, Ihsan / Jürges, Christopher / Juranic Lisnic, Vanda / L'Hernault, Anne / Rutkowski, Andrzej J / Prusty, Bhupesh K / Grothey, Arnhild / Milic, Andrea / Hennig, Thomas / Jonjic, Stipan / Friedel, Caroline C / Erhard, Florian / Dölken, Lars

PLoS pathogens

2023 Volume 19, Issue 5, Page(s) e1010992

Abstract: The genomes of both human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV) were first sequenced over 20 years ago. Similar to HCMV, the MCMV genome had initially been proposed to harbor ≈170 open reading frames (ORFs). More recently, omics ... ...

Abstract	The genomes of both human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV) were first sequenced over 20 years ago. Similar to HCMV, the MCMV genome had initially been proposed to harbor ≈170 open reading frames (ORFs). More recently, omics approaches revealed HCMV gene expression to be substantially more complex comprising several hundred viral ORFs. Here, we provide a state-of-the art reannotation of lytic MCMV gene expression based on integrative analysis of a large set of omics data. Our data reveal 365 viral transcription start sites (TiSS) that give rise to 380 and 454 viral transcripts and ORFs, respectively. The latter include >200 small ORFs, some of which represented the most highly expressed viral gene products. By combining TiSS profiling with metabolic RNA labelling and chemical nucleotide conversion sequencing (dSLAM-seq), we provide a detailed picture of the expression kinetics of viral transcription. This not only resulted in the identification of a novel MCMV immediate early transcript encoding the m166.5 ORF, which we termed ie4, but also revealed a group of well-expressed viral transcripts that are induced later than canonical true late genes and contain an initiator element (Inr) but no TATA- or TATT-box in their core promoters. We show that viral upstream ORFs (uORFs) tune gene expression of longer viral ORFs expressed in cis at translational level. Finally, we identify a truncated isoform of the viral NK-cell immune evasin m145 arising from a viral TiSS downstream of the canonical m145 mRNA. Despite being ≈5-fold more abundantly expressed than the canonical m145 protein it was not required for downregulating the NK cell ligand, MULT-I. In summary, our work will pave the way for future mechanistic studies on previously unknown cytomegalovirus gene products in an important virus animal model.
MeSH term(s)	Animals ; Mice ; Humans ; Muromegalovirus ; Cytomegalovirus/genetics ; Cytomegalovirus/metabolism ; Base Sequence ; Viral Proteins/genetics ; Viral Proteins/metabolism ; Open Reading Frames
Chemical Substances	Viral Proteins
Language	English
Publishing date	2023-05-12
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2205412-1
ISSN	1553-7374 ; 1553-7374
ISSN (online)	1553-7374
ISSN	1553-7374
DOI	10.1371/journal.ppat.1010992
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