The overall goals of this project are to understand how polyoma virus RNAs are made and regulated in infected cells and to use this virus as a model system to learn more general rules for mammalian RNA processing and function. We have discovered that much regulation of viral gene expression depends on the formation of double-stranded RNAs and their editing by the cellular dsRNA specific adenosine deaminase (ADAR1). Experiments completed during the previous funding period suggested that the long-sought trigger for the viral early-late switch might be the overlap and perhaps also the editing of the early and late polyadenylation signals. This represents a new mechanism of gene regulation. We also learned much new about nuclear responses to dsRNAs, including the involvement of a long nuclear-retained noncoding RNA (NEAT1) in the assembly of nuclear bodies called paraspeckles, which function in the retention of edited RNAs. We will extend these studies in three related aims. In the first, we will use new methods to better characterize RNA expression in the viral life cycle and will determine the nature and interplay of DNA sequence elements that promote regulated gene expression. In the second aim we will examine more closely the fate of polyoma dsRNAs in the nucleus, and the impact of cellular dsRNA response pathways including paraspeckles on infection. In the final aim we will carry out studies to learn more about how antisense RNA works in mammalian nuclei. This work will involve studying whether single DNA molecules can allow concurrent transcription in both directions as well as a new approach using small molecule-directed heterodimerization to alter the intranuclear proximity of DNA and RNA molecules.

Public Health Relevance

Mouse polyoma virus regulates the expression of its genes in an unusual manner, involving the generation of double-stranded RNA (dsRNA) molecules which play an important role in the viral life cycle. This proposal is to learn more about the underlying mechanisms of gene regulation in this system and to apply these lessons to achieve a fuller understanding of the function and fate of dsRNAs in mammalian cells.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA045382-27
Application #
8699149
Study Section
Virology - A Study Section (VIRA)
Program Officer
Read-Connole, Elizabeth Lee
Project Start
1987-07-01
Project End
2016-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
27
Fiscal Year
2014
Total Cost
$309,682
Indirect Cost
$106,880
Name
University of Connecticut
Department
Genetics
Type
Schools of Medicine
DUNS #
022254226
City
Farmington
State
CT
Country
United States
Zip Code
06030
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Chen, Ling-Ling; Carmichael, Gordon G (2009) Altered nuclear retention of mRNAs containing inverted repeats in human embryonic stem cells: functional role of a nuclear noncoding RNA. Mol Cell 35:467-78
Gu, Rui; Zhang, Zuo; DeCerbo, Joshua N et al. (2009) Gene regulation by sense-antisense overlap of polyadenylation signals. RNA 15:1154-63
Gu, R; Zhang, Z; Carmichael, G G (2006) How a small DNA virus uses dsRNA but not RNAi to regulate its life cycle. Cold Spring Harb Symp Quant Biol 71:293-9
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Kwon, Y G; Huang, H B; Desdouits, F et al. (1997) Characterization of the interaction between DARPP-32 and protein phosphatase 1 (PP-1): DARPP-32 peptides antagonize the interaction of PP-1 with binding proteins. Proc Natl Acad Sci U S A 94:3536-41

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