Transcriptional perturbations caused by SV40 large T antigen and its fragments
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE40567 | GEO | 2012/10/26
SECONDARY ACCESSION(S): PRJNA174316
REPOSITORIES: GEO
Similar Datasets
Project description:The small genome of polyomaviruses encodes a limited number of proteins that are highly dependent on interactions with host cell proteins for efficient viral replication. The SV40 large T antigen (LT) contains several discrete functional domains that contribute to the viral life cycle, including the DNA binding and helicase domains. In addition, the LT the C-terminal region is required for lytic infection in certain restrictive cell types. To understand how LT affects the host cell to facilitate viral replication, we expressed full-length or functional domains of LT in cells and identified interacting cellular proteins and performed expression profiling. LT perturbed the expression of p53 target genes and subsets of cell-cycle dependent genes regulated by the DREAM and the B-Myb-MuvB complexes. To examine transcriptome perturbations directly in human cells, we generated expression constructs of each LT fragment. Full length and truncated LT containing N-terminal HA and FLAG epitope tags were expressed from the pMSCV retroviral vector and introduced into U-2 OS cells. Total RNA was isolated from three biological replicate U-2 OS stable cell lines and gene expression was assayed on Affymetrix Human Gene U133 Plus 2.0 arrays.
2012-10-26 | E-GEOD-40567 | biostudies-arrayexpress
Project description:Polyomaviruses are small, circular dsDNA viruses that can cause cancer. Alternative splicing of polyomavirus early transcripts generates large and small tumor antigens (LT, ST) that play essential roles in viral replication and tumorigenesis. Some polyomaviruses also express middle tumor antigens (MTs) or alternate LT open reading frames (ALTOs), which are evolutionarily related but have distinct gene structures. MTs are a splice variant of the early transcript whereas ALTOs are overprinted on the second exon of the LT transcript in an alternate reading frame and are translated via an alternative start codon. Merkel cell polyomavirus (MCPyV), the only human polyomavirus that causes cancer, encodes an ALTO but its role in the viral lifecycle and tumorigenesis has remained elusive. In this MassIVE entry, we provide mass spectrometry data and result files associated with a TurboID experiment to identify human and viral proteins that bind to ALTO.
2024-07-26 | MSV000095457 | MassIVE
Project description:Polyomaviruses are a family of small DNA tumor viruses that includes several pathogenic human members, including Merkel cell polyomavirus, BK virus and JC virus. As is characteristic of DNA tumor viruses, gene expression in polyomaviruses is temporally regulated into an early phase, consisting of the viral regulatory proteins, and a late phase, consisting of the viral structural proteins. Previously, the late transcripts expressed by the prototypic polyomavirus simian virus 40 (SV40) were reported to contain several adenosines bearing methyl groups at the N6 position (m6A), although the precise location of these m6A residues, and their phenotypic effects, have not been investigated. Here, we first demonstrate that overexpression of the key m6A reader protein YTHDF2 induces more rapid viral replication, and larger viral plaques, in SV40 infected BSC40 cells, while mutational inactivation of the endogenous YTHDF2 gene, or the m6A methyltransferase METTL3, has the opposite effect, thus suggesting a positive role for m6A in the regulation of SV40 gene expression. To directly test this hypothesis, we mapped sites of m6A addition on SV40 transcripts and identified two m6A sites on the viral early transcripts and eleven m6A sites on the late mRNAs. Using synonymous mutations, we inactivated the majority of the m6A sites on the SV40 late mRNAs and observed that the resultant viral mutant replicated more slowly than wild type SV40. Alternative splicing of SV40 late mRNAs was unaffected by the reduction in m6A residues and our data instead suggest that m6A enhances the translation of viral late transcripts. Together, these data argue that the addition of m6A residues to the late transcripts encoded by SV40 plays an important role in enhancing viral gene expression and, hence, replication.
2018-02-08 | GSE106698 | GEO
Project description:Myb-MuvB (MMB)/dREAM is a nine subunit complex first described in Drosophila as a repressor of transcription, dependent upon E2F2 and the RBFs. Myb, an integral member of MMB, curiously plays no role in the silencing of the test genes previously analyzed. Moreover, Myb plays an activating role in DNA replication in Drosophila egg chamber follicle cells. The essential functions for Myb are executed as part of MMB. This duality of function lead to the hypothesis that MMB, which contains both known activator and repressor proteins, might function as part of a switching mechanism that is dependent upon DNA sites and developmental context. Keywords: Drosophila Myb-MuvB/dREAM, ChIP-chip
2007-11-14 | GSE9087 | GEO
Project description:Polyomaviruses are small, circular dsDNA viruses that can cause cancer. Alternative splicing of polyomavirus early transcripts generates large and small tumor antigens (LT, ST) that play essential roles in viral replication and tumorigenesis. Some polyomaviruses also express middle tumor antigens (MTs) or alternate LT open reading frames (ALTOs), which are evolutionarily related but have distinct gene structures. MTs are a splice variant of the early transcript whereas ALTOs are overprinted on the second exon of the LT transcript in an alternate reading frame and are translated via an alternative start codon. Merkel cell polyomavirus (MCPyV), the only human polyomavirus that causes cancer, encodes an ALTO but its role in the viral lifecycle and tumorigenesis has remained elusive. Here, we performed bulk RNA sequencing in MKL-2 cells, a Merkel cell carcinoma cell line, expressing MCPyV ALTO or RFP as a negative control via a doxycycline inducible lentiviral vector. The goal of the study was to identify Differentially Expressed Genes and pathways activated or inhibited via Gene Set Enrichment Analysis.
2024-09-01 | GSE273479 | GEO
Project description:Polyomaviruses (PyVs) cause diverse diseases in a variety of mammalian hosts. During the life cycle, PyVs recruit nuclear host factors to viral genomes to facilitate replication and transcription. While host factors involved in DNA replication, DNA damage sensing and repair, and cell cycle regulation have been observed to bind PyV DNA, the complete set of viral and host proteins comprising the PyV replisome remains incompletely characterized. Here, the iPOND-MS technique (Isolation of Proteins on Nascent DNA coupled with Mass Spectrometry) was used to identify the proteome bound to murine PyV (MuPyV) DNA immediately following synthesis and two hours post-synthesis. Several novel MuPyV DNA interactors were identified on newly synthesized viral DNA (vDNA), including MCM complex members, DNA primase, DNA polymerase alpha, DNA ligase, and replication factor C. Though displaying partial overlap, the set of host and viral proteins bound to MuPyV DNA two hours post-synthesis lacked many of the replication proteins found on newly synthesized vDNA. These data help distinguish between the host factors critical for MuPyV DNA replication and those involved in downstream processing.
2024-10-07 | PXD055210 | Pride
Project description:The SV40 large (LT) and small (st) antigens are produced from a single alternatively spliced pre-mRNA, that when co-expressed, transform a variety of cells in vitro and in vivo. However, 17kT, a relatively uncharacterized third protein that is co-linear with LT for the first 131 amino acids, is also produced from the early viral pre-mRNA by removal of an additional intron from the LT transcript. Here we report a line of transgenic mice expressing a liver-specific dox-inducible viral transcript that fails to yield any detectable LT protein, yet produces abundant 17kT. Comparative analysis of livers of transgenic mice expressing either 17kT or LT demonstrates that while 17kT is a potent stimulator of cell proliferation, it is ineffective at inducing liver tumor development, due in part, to the failure of 17kT to effectively induce the expression of growth regulators and reactivate expression of imprinted and developmentally regulated hepatic genes. These studies highlight key functional differences between LT and 17kT in their ability to transform quiescent primary epithelial cells in vivo, and demonstrate how specific functional domains within LT impact cell-specific gene expression to promote oncogenesis. Keywords: genetic modification, SV40, 17kT, LT, Rb, p53, hepatocellular carcinoma, hepatic hyperplasia, differentiation.
2010-04-01 | GSE5242 | GEO
Project description:The SV40 large (LT) and small (st) antigens are produced from a single alternatively spliced pre-mRNA, that when co-expressed, transform a variety of cells in vitro and in vivo. However, 17kT, a relatively uncharacterized third protein that is co-linear with LT for the first 131 amino acids, is also produced from the early viral pre-mRNA by removal of an additional intron from the LT transcript. Here we report a line of transgenic mice expressing a liver-specific dox-inducible viral transcript that fails to yield any detectable LT protein, yet produces abundant 17kT. Comparative analysis of livers of transgenic mice expressing either 17kT or LT demonstrates that while 17kT is a potent stimulator of cell proliferation, it is ineffective at inducing liver tumor development, due in part, to the failure of 17kT to effectively induce the expression of growth regulators and reactivate expression of imprinted and developmentally regulated hepatic genes. These studies highlight key functional differences between LT and 17kT in their ability to transform quiescent primary epithelial cells in vivo, and demonstrate how specific functional domains within LT impact cell-specific gene expression to promote oncogenesis. Keywords: genetic modification, SV40, 17kT, LT, Rb, p53, hepatocellular carcinoma, hepatic hyperplasia, differentiation. Affymetrix mouse genome 430 A and B platform was used, single measurements for wild-type and transgenic animals.
2010-03-31 | E-GEOD-5242 | biostudies-arrayexpress
Project description:The mammalian Retinoblastoma (RB) family including pRB, p107, and p130 represses E2F target genes through mechanisms that are not fully understood. In D. melanogaster, RB-dependent repression is mediated in part by the multisubunit protein complex Drosophila RBF, E2F, and Myb (dREAM) that contains homologs of the C. elegans synthetic multivulva class B (synMuvB) gene products. Using an integrated approach combining proteomics, genomics, and bioinformatic analyses, we identified a p130 complex termed DP, RB-like, E2F, and MuvB (DREAM) that contains mammalian homologs of synMuvB proteins LIN-9, LIN-37, LIN-52, LIN-54, and LIN-53/RBBP4. DREAM bound to more than 800 human promoters in G0 and was required for repression of E2F target genes. In S phase, MuvB proteins dissociated from p130 and formed a distinct submodule that bound MYB. This work reveals an evolutionarily conserved multisubunit protein complex that contains p130 and E2F4, but not pRB, and mediates the repression of cell cycle-dependent genes in quiescence. Experiment Overall Design: Gene expression during the cell cycle in T98G cells. Cells were serum starved for 72 hours to induce G0 and then restimuated to enter cell cycle by serum addition. BrdU and FACS was done simultaneously to confirm their cell cycle phase.
2008-06-16 | E-GEOD-8537 | biostudies-arrayexpress