This competitive renewal application continues to focus on the early stages of retroviral replication. The overarching goal of this application is to uncover critical cell-virus interactions that determine the success or failure of retroviral gene expression in human cells, as modulated by epigenetic silencing. It was established decades ago that the prototype avian sarcoma virus (ASV) DNA provirus is subject to epigenetic silencing in mammalian cells, and this system will continue to be studied as a model. The applicants have discovered that epigenetic silencing of ASV can be mediated by the rapid engagement of an antiviral host adaptor protein, Daxx, which recruits the cellular epigenetic silencing machinery. Furthermore, through development and implementation of an innovative, robust, and comprehensive siRNA-based screen, a number of novel and expected human host cell factors that play a role in this response were identified. These findings have opened up the exciting new areas of investigation to be pursued in this competing renewal.
In Aim 1, the molecular mechanisms that govern Daxx function as an antiviral factor will be determined. Results of these studies will provide valuable insight into general intrinsic host defense mechanisms.
In Aim 2 mechanistic details for the function of several novel factors identified by siRNA screening will be studied, with a focus on those likely to engage viral DNA sequences. The hypotheses that specific viral DNA binding factors either participate in nucleation of silencing factors or act as a transcriptional barrier, will be tested.
Aim 3 will exploit new comprehensive and systematic approaches to distinguish the physical and functional epigenetic features of silent proviruses that are established soon after infection (initiation) and after long-term passage (maintenance). Specific hypotheses concerning the roles of antiviral factors, and the interplay of proviral and cellular chromatin, in silencing will be considered. Epigenetic mechanisms have an essential role in the regulation of gene expression during development, in differentiated adult tissues, and in human disease, including cancer. Consequently, results from the proposed research will be highly relevant to virology, as well as the cellular biology of normal and cancer cells.

Public Health Relevance

Cells employ epigenetic silencing mechanisms to repress expression of retroviral genes, and the proposed research will increase our understanding of how such silencing is initiated and maintained in human cells. The expression of cellular genes is also regulated by epigenetic mechanisms during development, and failure or inappropriate silencing can lead to developmental errors and diseases such as cancer. These studies will help to uncover new strategies by which appropriate silencing may be restored, and disease prevented or treated.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA071515-18
Application #
8606819
Study Section
Virology - A Study Section (VIRA)
Program Officer
Daschner, Phillip J
Project Start
1996-09-30
Project End
2016-11-30
Budget Start
2013-12-01
Budget End
2014-11-30
Support Year
18
Fiscal Year
2014
Total Cost
$452,394
Indirect Cost
$198,952
Name
Research Institute of Fox Chase Cancer Center
Department
Type
DUNS #
064367329
City
Philadelphia
State
PA
Country
United States
Zip Code
19111
Balasubramanian, Sangeetha; Rajagopalan, Muthukumaran; Bojja, Ravi Shankar et al. (2017) The conformational feasibility for the formation of reaching dimer in ASV and HIV integrase: a molecular dynamics study. J Biomol Struct Dyn 35:3469-3485
Andrake, Mark D; Skalka, Anna Marie (2015) Retroviral Integrase: Then and Now. Annu Rev Virol 2:241-64
Peretz, Yuval; Wu, Hong; Patel, Shayan et al. (2015) Inhibitor of DNA Binding 4 (ID4) is highly expressed in human melanoma tissues and may function to restrict normal differentiation of melanoma cells. PLoS One 10:e0116839
Skala, Anna Marie (2014) Retroviral DNA Transposition: Themes and Variations. Microbiol Spectr 2:
Skalka, Anna Marie (2014) Retroviral DNA Transposition: Themes and Variations. Microbiol Spectr 2:MDNA300052014
Poleshko, Andrey; Kossenkov, Andrew V; Shalginskikh, Natalia et al. (2014) Human factors and pathways essential for mediating epigenetic gene silencing. Epigenetics 9:1280-9
Haugh, Kelsey A; Shalginskikh, Natalia; Nogusa, Shoko et al. (2014) The interferon-inducible antiviral protein Daxx is not essential for interferon-mediated protection against avian sarcoma virus. Virol J 11:100
Poleshko, Andrey; Katz, Richard A (2014) Specifying peripheral heterochromatin during nuclear lamina reassembly. Nucleus 5:32-9
Poleshko, Andrey; Mansfield, Katelyn M; Burlingame, Caroline C et al. (2013) The human protein PRR14 tethers heterochromatin to the nuclear lamina during interphase and mitotic exit. Cell Rep 5:292-301
Bojja, Ravi Shankar; Andrake, Mark D; Merkel, George et al. (2013) Architecture and assembly of HIV integrase multimers in the absence of DNA substrates. J Biol Chem 288:7373-86

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