JC virus agnoprotein enhances large T antigen binding to the origin of viral DNA replication: Evidence for its involvement in viral DNA replication

Sarıbaş, A. Sami; White, Martyn K.; Safak, Mahmut

doi:10.1016/j.virol.2012.06.017

Cited by 29 publications

(56 citation statements)

References 84 publications

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“…This type of behavior by the L29A mutant suggests that agnoprotein may shuttle between the nucleus and cytoplasm to fulfill its regulatory functions during the viral replication cycle. Consistent with this concept, we have recently demonstrated that agnoprotein enhances the binding activity of JCV large T antigen to the origin of viral DNA replication (4). Moreover, consistent with the nuclear localization of the L29A mutant, we have recently demonstrated that various ␣-helix domain mutants of agnoprotein also showed defects in splicing of 6), and was subjected to RT-PCR using specific primers, as described in Materials and Methods.…”

Section: Discussionsupporting

confidence: 63%

“…Agnoprotein is a cytoplasmic protein, with high concentrations accumulating in the perinuclear region of infected cells, but a small portion of the protein is also consistently detected in the nucleus, indicating a possible nuclear function for agnoprotein. To support this notion in functional assays, we demonstrated that agnoprotein enhances the DNA binding activity of LT-Ag to the viral origin (Ori) without itself directly interacting with DNA (4). We also demonstrated that agnoproteins of BKV, SV40, and JCV form highly stable, SDS-resistant homodimers and oligomers (7,18).…”

mentioning

confidence: 84%

“…It is also interesting to note that all three Phe residues of agnoprotein (Phe31, Phe35, and Phe39) localize to its hydrophobic region. The functional importance of these Phe residues has recently been evaluated by genetic and biochemical approaches, and it was demonstrated that they positively influence viral DNA replication (1,4) and play regulatory roles in the perinuclear distribution of agnoprotein in infected cells (4,5). Agnoprotein possesses only one highly conserved Cys residue, which is located right after the hydrophobic region of the protein (Cys40).…”

mentioning

confidence: 99%

“…JCV agnoprotein was previously shown to interact with a number of cellular and viral proteins, including YB-1 (11), p53 (12), FEZ1 and HP1-␣ (13), adaptor protein complex 3 (14), JCV small t antigen (Sm t-Ag) (15), and JCV large T antigen (LT-Ag) (16), and has been implicated in various aspects of the JCV life cycle, including viral replication (4,17), viral transcription (11), functioning as a viroporin (14,18), encapsidation (19), and interfering with exocytosis (20). In addition, this protein deregulates cell cycle progression, where cells stably expressing agnoprotein largely accumulate at the G 2 /M phase of the cell cycle (12).…”

mentioning

confidence: 99%

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Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain

Coric

Sarıbaş

Abou‐Gharbia

et al. 2014

J Virol

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Agnoprotein is a small multifunctional regulatory protein required for sustaining the productive replication of JC virus (JCV). It is a mostly cytoplasmic protein localizing in the perinuclear area and forms highly stable dimers/oligomers through a Leu/Ile/ Phe-rich domain. There have been no three-dimensional structural data available for agnoprotein due to difficulties associated with the dynamic conversion from monomers to oligomers. Here, we report the first nuclear magnetic resonance (NMR) structure of a synthetic agnoprotein peptide spanning amino acids Thr17 to Glu55 where Lys23 to Phe39 encompassing the Leu/Ile/ Phe-rich domain forms an amphipathic ␣-helix. On the basis of these structural data, a number of Ala substitution mutations were made to investigate the role of the ␣-helix in the structure and function of agnoprotein. Single L29A and L36A mutations exhibited a significant negative effect on both protein stability and viral replication, whereas the L32A mutation did not. In addition, the L29A mutant displayed a highly nuclear localization pattern, in contrast to the pattern for the wild type (WT). Interestingly, a triple mutant, the L29A؉L32A؉L36A mutant, yielded no detectable agnoprotein expression, and the replication of this JCV mutant was significantly reduced, suggesting that Leu29 and Leu36 are located at the dimer interface, contributing to the structure and stability of agnoprotein. Two other single mutations, L33A and E34A, did not perturb agnoprotein stability as drastically as that observed with the L29A and L36A mutations, but they negatively affected viral replication, suggesting that the role of these residues is functional rather than structural. Thus, the agnoprotein dimerization domain can be targeted for the development of novel drugs active against JCV infection. IMPORTANCEAgnoprotein is a small regulatory protein of JC virus (JCV) and is required for the successful completion of the viral replication cycle. It forms highly stable dimers and oligomers through its hydrophobic (Leu/Ile/Phe-rich) domain, which has been shown to play essential roles in the stability and function of the protein. In this work, the Leu/Ile/Phe-rich domain has been further characterized by NMR studies using an agnoprotein peptide spanning amino acids T17 to Q54. Those studies revealed that the dimerization domain of the protein forms an amphipathic ␣-helix. Subsequent NMR structure-based mutational analysis of the region highlighted the critical importance of certain amino acids within the ␣-helix for the stability and function of agnoprotein. In conclusion, this study provides a solid foundation for developing effective therapeutic approaches against the dimerization domain of the protein to inhibit its critical roles in JCV infection.

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Section: Discussionsupporting

confidence: 63%

mentioning

confidence: 84%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain

Coric

Sarıbaş

Abou‐Gharbia

et al. 2014

J Virol

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“…Phenylalanine residues are known to have critical roles in protein-protein interaction, protein folding, and stability. Mutation of all the 3 phenylalanine residues present in the agnoprotein resulted in inefficient replication of mutant virus (Saribas et al 2012). …”

Section: Dna Replicationmentioning

confidence: 99%

Entry, infection, replication, and egress of human polyomaviruses: an update

Bhattacharjee

Chattaraj

2017

Can. J. Microbiol.

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Polyomaviruses (PyVs), belonging to the family Polyomaviridae, are a group of small, nonenveloped, double-stranded, circular DNA viruses widely distributed in the vertebrates. PyVs cause no apparent disease in adult laboratory mice but cause a wide variety of tumors when artificially inoculated into neonates or semipermissive animals. A few human PyVs, such as BK, JC, and Merkel cell PyVs, have been unequivocally linked to pathogenesis under conditions of immunosuppression. Infection is thought to occur early in life and persists for the lifespan of the host. Over evolutionary time scales, it appears that PyVs have slowly co-evolved with specific host animal lineages. Host cell surface glycoproteins and glycolipids seem to play a decisive role in the entry stage of viral infection and in channeling the virions to specific intracellular membrane-bound compartments and ultimately to the nucleus, where the genomes are replicated and packaged for release. Therefore the transport of the infecting virion or viral genome to this site of multiplication is an essential process in productive viral infection as well as in latent infection and transformation. This review summarizes the major findings related to the characterization of the nature of the interactions between PyV and host protein and their impact in host cell invasion.

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Nuclear Magnetic Resonance Structure of the Human Polyoma JC Virus Agnoprotein

Coric

Abou‐Gharbia

et al. 2017

J of Cellular Biochemistry

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Agnoprotein is an important regulatory protein of the human polyoma JC virus (JCV) and plays critical roles during the viral replication cycle. It forms highly stable dimers and oligomers through its Leu/Ile/Phe-rich domain, which is important for the stability and function of the protein. We recently resolved the partial 3D structure of this protein by NMR using a synthetic peptide encompassing amino acids Thr17 to Gln52, where the Leu/Ile/Phe- rich domain was found to adopt a major alpha-helix conformation spanning amino acids 23 to 39. Here, we report the resolution of the 3D structure of full-length JCV agnoprotein by NMR, which not only confirmed the existence of the previously reported major α-helix domain at the same position but also revealed the presence of an additional minor α-helix region spanning amino acid residues Leu6 to Ala10. The remaining regions of the protein adopt an intrinsically unstructured conformation.

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JC virus agnoprotein enhances large T antigen binding to the origin of viral DNA replication: Evidence for its involvement in viral DNA replication

Cited by 29 publications

References 84 publications

Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain

Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain

Entry, infection, replication, and egress of human polyomaviruses: an update

Nuclear Magnetic Resonance Structure of the Human Polyoma JC Virus Agnoprotein

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