JC virus (JCV) is an etiological agent of Progressive Multifocal Leukoencephalopathy (PML), a fatal demyelinating disease of the Central Nervous System (CNS), which is frequently seen in patients with underlying immunosuppressive conditions, including leukemia, lymphomas and AIDS. PML in the era of AIDS epidemic dramatically increased which makes it an AIDS defining condition. JCV establishes a sub-clinical latent infection in the body but upon reactivation, it enters the brain;and lytically and abortively infects oligodendrocytes (myelin producing cells) and astrocytes respectively. This infection results in an extensive myelin loss in CNS, which is the characteristic histopathological landmark of PML. This is a revised competitive renewal R01 grant application, entitled "Role of agnoprotein in JC virus life cycle". During the course of this funding period, we established that agnoprotein of JCV plays important regulatory roles in JCV life cycle through molecular interactions with a cellular transcription factor, YB-1 and the viral regulatory protein, large T antigen. It deregulates cell cycle progression, in which agnoprotein positive cells largely accumulate at G2/M phase transition. We also demonstrated the involvement of the coding region of agnoprotein in regulation of JCV life cycle by deletion analysis. In this regard, we showed that agnoprotein- coding region contains important cis-acting DNA elements to which specific transcription factors bind and contribute the viral life cycle. Furthermore, analysis of the PKC phosphorylation sites of this protein by mutagenesis revealed the fact that phosphorylation plays a critical role in the function of agnoprotein, because none of the phosphorylation mutants (Ser7, Ser11 and Thr21 to Ala) was able to continue viral replication cycle due to a limited replication and empty capsid formation. Moreover, our recent findings also showed that phosphorylated form of agnoprotein is targeted by a Ser/Thr phosphatase, PP2A, for dephosphorylation and this can be inhibited by JCV small t antigen, suggesting a functional ternary complex formation between these three proteins. These findings collectively have provided us a strong rationale to further study the functions of agnoprotein and led us to hypothesize that agnoprotein plays critical regulatory roles in JCV virion biogenesis and therefore in the progression of PML. As such, the understanding of the molecular mechanisms in which agnoprotein is involved in JCV replication is central to unravel the molecular mechanisms that are critical for the disease progression so that we would be able to develop effective therapeutic strategies against this disease. Thus, to further examine our hypothesis, we propose to i) investigate the molecular mechanisms by which agnoprotein regulates both the virion formation and large T antigen-mediated viral DNA replication;and ii) examine the impact of both PP2A and Sm t-Ag on agnoprotein functions during this process.

Public Health Relevance

Progressive multifocal leukoencephalopathy (PML), a white mater disease of the central nervous system, is caused by a human neurotropic polyomavirus, JC virus (JCV). This disease mostly affects immunocompromised patients with underlying disorders such as lymphoproliferative and myeloproliferative diseases and acquired immunodeficiency syndrome, (AIDS). In this research project, we are proposing experiments to understand the role of one of JCV regulatory proteins, agnoprotein in viral biogenesis, which may allow us to design effective therapeutic strategies to curb the disease in affected individuals.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS043108-10
Application #
8302988
Study Section
NeuroAIDS and other End-Organ Diseases Study Section (NAED)
Program Officer
Wong, May
Project Start
2001-12-01
Project End
2014-07-31
Budget Start
2012-08-01
Budget End
2014-07-31
Support Year
10
Fiscal Year
2012
Total Cost
$367,500
Indirect Cost
$122,500
Name
Temple University
Department
Neurosciences
Type
Schools of Medicine
DUNS #
057123192
City
Philadelphia
State
PA
Country
United States
Zip Code
19122
Saribas, A Sami; Mun, Sarah; Johnson, Jaslyn et al. (2014) Human polyoma JC virus minor capsid proteins, VP2 and VP3, enhance large T antigen binding to the origin of viral DNA replication: evidence for their involvement in regulation of the viral DNA replication. Virology 449:1-16
Coric, Pascale; Saribas, A Sami; Abou-Gharbia, Magid et al. (2014) Nuclear magnetic resonance structure revealed that the human polyomavirus JC virus agnoprotein contains an ?-helix encompassing the Leu/Ile/Phe-rich domain. J Virol 88:6556-75
Sami Saribas, A; Abou-Gharbia, Magid; Childers, Wayne et al. (2013) Essential roles of Leu/Ile/Phe-rich domain of JC virus agnoprotein in dimer/oligomer formation, protein stability and splicing of viral transcripts. Virology 443:161-76
Saribas, A Sami; Arachea, Buenafe T; White, Martyn K et al. (2011) Human polyomavirus JC small regulatory agnoprotein forms highly stable dimers and oligomers: implications for their roles in agnoprotein function. Virology 420:51-65
Khalili, Kamel; Sariyer, Ilker Kudret; Safak, Mahmut (2008) Small tumor antigen of polyomaviruses: role in viral life cycle and cell transformation. J Cell Physiol 215:309-19
Romagnoli, Luca; Sariyer, Ilker K; Tung, Jacqueline et al. (2008) Early growth response-1 protein is induced by JC virus infection and binds and regulates the JC virus promoter. Virology 375:331-41
Staniszewska, Izabela; Sariyer, Ilker K; Lecht, Shimon et al. (2008) Integrin alpha9 beta1 is a receptor for nerve growth factor and other neurotrophins. J Cell Sci 121:504-13
Sariyer, Ilker K; Khalili, Kamel; Safak, Mahmut (2008) Dephosphorylation of JC virus agnoprotein by protein phosphatase 2A: inhibition by small t antigen. Virology 375:464-79