Nucleosome sliding, histone modifications, and the epigenetic regulation of the Simian Virus 40 life cycle

ABSTRACTSimian virus 40 (SV40) exists as chromatin throughout its life cycle and undergoes typical epigenetic regulation mediated by changes in nucleosome location and associated histone modifications. In order to investigate the role of epigenetic regulation during the encapsidation of late-stage minichromosomes into virions, we mapped the locations of nucleosomes containing acetylated or methylated lysines in the histone tails of H3 and H4 present in the chromatin from 48-h-postinfection minichromosomes and disrupted virions. In minichromosomes obtained late in infection, nucleosomes were found carrying various histone modifications primarily in the regulatory region, with a major nucleosome located within the enhancer and other nucleosomes at the early and late transcriptional start sites. The nucleosome found in the enhancer would be expected to repress early transcription by blocking access to part of the SP1 binding sites and the left side of the enhancer in late-stage minichromosomes while also allowing late transcription. In chromatin from virions, the principal nucleosome located in the enhancer was shifted ∼70 bases in the late direction from what was found in minichromosomes, and the level of modified histones was increased throughout the genome. The shifting of the enhancer-associated nucleosome to the late side would effectively serve as a switch to relieve the repression of early transcription found in late minichromosomes while likely also repressing late transcription by blocking access to necessary regulatory sequences. This epigenetic switch appeared to occur during the final stage of virion formation.IMPORTANCEFor a virus to complete infection, it must produce a new virus particle in which the genome is able to support a new infection. This is particularly important for viruses like simian virus 40 (SV40), which exist as chromatin throughout their life cycles, since chromatin structure plays a major role in the regulation of the life cycle. In order to determine the role of SV40 chromatin structure late in infection, we mapped the locations of nucleosomes and their histone tail modifications in SV40 minichromosomes and in the SV40 chromatin found in virions using chromatin immunoprecipitation-DNA sequencing (ChIP-Seq). We have identified a novel viral transcriptional control mechanism in which a nucleosome found in the regulatory region of the SV40 minichromosome is directed to slide during the formation of the virus particle, exposing transcription factor binding sites required for early transcription that were previously blocked by the presence of the nucleosome.

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Directed Nucleosome Sliding in SV40 Minichromosomes During the Formation of the Virus Particle Exposes DNA Sequences Required for Early Transcription

10.1101/426452 ◽

2018 ◽

Cited By ~ 2

Author(s):

Meera Ajeet Kumar ◽

Karine Kasti ◽

Lata Balakrishnan ◽

Barry Milavetz

Keyword(s):

Histone Modifications ◽

Epigenetic Regulation ◽

Dna Sequences ◽

Late Stage ◽

Regulatory Region ◽

Simian Virus 40 ◽

Simian Virus ◽

Regulatory Sequences ◽

Late Transcription ◽

Early Transcription

AbstractSimian Virus 40 (SV40) exists as chromatin throughout its life cycle, and undergoes typical epigenetic regulation mediated by changes in nucleosome location and associated histone modifications. In order to investigate the role of epigenetic regulation during the encapsidation of late stage minichromosomes into virions, we have mapped the location of nucleosomes containing acetylated or methylated lysines in the histone tails of H3 and H4 present in the chromatin from 48-hour post-infection minichromosomes and disrupted virions. In minichromosomes obtained late in infection, nucleosomes were found carrying various histone modifications primarily in the regulatory region with a major nucleosome located within the enhancer and other nucleosomes at the early and late transcriptional start sites. The nucleosome found in the enhancer would be expected to repress early transcription by blocking access to part of the SP1 binding sites and the left side of the enhancer in late stage minichromosomes while also allowing late transcription. In chromatin from virions, the principal nucleosome located in the enhancer was shifted ~ 70 bases in the late direction from what was found in minichromosomes and the level of modified histones was increased throughout the genome. The shifting of the enhancer-associated nucleosome to the late side would effectively serve as a switch to relieve the repression of early transcription found in late minichromosomes while likely also repressing late transcription by blocking access to necessary regulatory sequences. This epigenetic switch appeared to occur during the final stage of virion formation.

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The Abundant Nuclear Enzyme PARP Participates in the Life Cycle of Simian Virus 40 and Is Stimulated by Minor Capsid Protein VP3

Journal of Virology ◽

10.1128/jvi.77.7.4273-4282.2003 ◽

2003 ◽

Vol 77 (7) ◽

pp. 4273-4282 ◽

Cited By ~ 25

Author(s):

Ariela Gordon-Shaag ◽

Yael Yosef ◽

Mahmoud Abd El-Latif ◽

Ariella Oppenheim

Keyword(s):

Amino Acids ◽

Life Cycle ◽

Capsid Protein ◽

Membrane Integrity ◽

Simian Virus 40 ◽

Simian Virus ◽

Capsid Proteins ◽

Dependent Manner ◽

Minor Capsid Protein ◽

Stimulation Of

ABSTRACT The abundant nuclear enzyme poly(ADP-ribose) polymerase (PARP) functions in DNA damage surveillance and repair and at the decision between apoptosis and necrosis. Here we show that PARP binds to simian virus 40 (SV40) capsid proteins VP1 and VP3. Furthermore, its enzymatic activity is stimulated by VP3 but not by VP1. Experiments with purified mutant proteins demonstrated that the PARP binding domain in VP3 is localized to the 35 carboxy-terminal amino acids, while a larger peptide of 49 amino acids was required for full stimulation of its activity. The addition of 3-aminobenzamide (3-AB), a known competitive inhibitor of PARP, demonstrated that PARP participates in the SV40 life cycle. The titer of SV40 propagated on CV-1 cells was reduced by 3-AB in a dose-dependent manner. Additional experiments showed that 3-AB did not affect viral DNA replication or capsid protein production. PARP did not modify the viral capsid proteins in in vitro poly(ADP-ribosylation) assays, implying that it does not affect SV40 infectivity. On the other hand, it greatly reduced the magnitude of the host cytopathic effects, a hallmark of SV40 infection. Additional experiments suggested that the stimulation of PARP activity by VP3 leads the infected cell to a necrotic pathway, characterized by the loss of membrane integrity, thus facilitating the release of mature SV40 virions from the cells. Our studies identified a novel function of the minor capsid protein VP3 in the recruitment of PARP for the SV40 lytic process.

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Intracellular forms of simian virus 40 nucleoprotein complexes. III. Study of histone modifications.

Journal of Virology ◽

10.1128/jvi.36.2.353-360.1980 ◽

1980 ◽

Vol 36 (2) ◽

pp. 353-360 ◽

Cited By ~ 13

Author(s):

M Coca-Prados ◽

G Vidali ◽

M T Hsu

Keyword(s):

Histone Modifications ◽

Simian Virus 40 ◽

Simian Virus ◽

Nucleoprotein Complexes

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Role of Simian Virus 40 Vp1 Cysteines in Virion Infectivity

Journal of Virology ◽

10.1128/jvi.74.23.11388-11393.2000 ◽

2000 ◽

Vol 74 (23) ◽

pp. 11388-11393 ◽

Cited By ~ 25

Author(s):

Peggy P. Li ◽

Akira Nakanishi ◽

Mary A. Tran ◽

Adler M. Salazar ◽

Robert C. Liddington ◽

...

Keyword(s):

Life Cycle ◽

Disulfide Bonds ◽

Three Dimensional ◽

Simian Virus 40 ◽

Experimental System ◽

Simian Virus ◽

Dimensional Structure ◽

Wild Type

ABSTRACT We have developed a new nonoverlapping infectious viral genome (NO-SV40) in order to facilitate structure-based analysis of the simian virus 40 (SV40) life cycle. We first tested the role of cysteine residues in the formation of infectious virions by individually mutating the seven cysteines in the major capsid protein, Vp1. All seven cysteine mutants—C9A, C49A, C87A, C104A, C207S, C254A, and C267L—retained viability. In the crystal structure of SV40, disulfide bridges are formed between certain Cys104 residues on neighboring pentamers. However, our results show that none of these disulfide bonds are required for virion infectivity in culture. We also introduced five different mutations into Cys254, the most strictly conserved cysteine across the polyomavirus family. We found that C254L, C254S, C254G, C254Q, and C254R mutants all showed greatly reduced (around 100,000-fold) plaque-forming ability. These mutants had no apparent defect in viral DNA replication. Mutant Vp1's, as well as wild-type Vp2/3, were mostly localized in the nucleus. Further analysis of the C254L mutant revealed that the mutant Vp1 was able to form pentamers in vitro. DNase I-resistant virion-like particles were present in NO-SV40-C254L-transfected cell lysate, but at about 1/18 the amount in wild-type-transfected lysate. An examination of the three-dimensional structure reveals that Cys254 is buried near the surface of Vp1, so that it cannot form disulfide bonds, and is not involved in intrapentamer interactions, consistent with the normal pentamer formation by the C254L mutant. It is, however, located at a critical junction between three pentamers, on a conserved loop (G2H) that packs against the dual interpentamer Ca2+-binding sites and the invading C-terminal helix of an adjacent pentamer. The substitution by the larger side chains is predicted to cause a localized shift in the G2H loop, which may disrupt Ca2+ ion coordination and the packing of the invading helix, consistent with the defect in virion assembly. Our experimental system thus allows dissection of structure-function relationships during the distinct steps of the SV40 life cycle.

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