Characterizing unique virus-host interactions is key to understanding pathogenesis and developing therapeutics to block the virus life cycle. Because of their intimate associations with host cells, viruses have also been exploited as tools for studying many basic cellular processes, including RNA processing. For retroviruses, control of splicing and polyadenylation are important aspects of the replication cycle. Splicing of retroviral primary transcripts must be controlled since high levels of unspliced RNA are needed as mRNA, and for incorporation as genomes into progeny virions. Polyadenylation control is important because failure to use the viral polyadenylation site results in read-through transcripts that extend into downstream genomic sequences;this is the basis for oncogenic transformation and the ability of retroviruses to acquire host cell sequences through oncogene capture. We are studying viral cis elements and host trans-acting factors required for proper RNA processing and replication of Rous sarcoma virus (RSV);paradigms established for RSV RNA processing control will provide insights into host cell RNA processing regulation. Our work focuses in part on a novel RNA processing control element, the negative regulator of splicing (NRS), that contributes to the accumulation of genome-length RNA by acting as a pseudo 5'ss to repress splicing. We will continue investigation of factors required for novel snRNP interactions with the NRS and their role in splicing control. Splicing and polyadenylation are coupled in the cells that RSV infects, raising the question of how viral RNAs are efficiently polyadenylated. We will continue examining a novel coupling mechanism whereby the NRS, in addition to splicing control, ensures the proper polyadenylation of genome-length RNA that do not benefit from splicing. RSV also harbors a site-specific splicing regulator, the suppressor of src splicing (SSS), that controls src splicing and is required for proper replication. We will identify host factors required for SSS activity and explore the mechanism by which it blocks splicing. Through this work we will enhance our understanding of control of critical RNA processing steps required for viral replication and pathogenesis, and could lead to new therapeutics designed to block replication. The RSV system also provides a powerful tool to dissect novel cellular mechanisms of RNA processing regulation.

Public Health Relevance

Retroviruses are major animal pathogens that cause considerable human disease, including immunodeficiencies and cancer. Viral replication requires unconventional use of the machines inside cells that make viral RNA. Understanding how these agents interact with manipulate the host cell RNA processing machinery could lead to new therapeutics designed to block replication and teach us novel ways in which the host machines work.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA078709-14
Application #
8284173
Study Section
Virology - A Study Section (VIRA)
Program Officer
Read-Connole, Elizabeth Lee
Project Start
1998-08-14
Project End
2014-06-30
Budget Start
2012-07-01
Budget End
2014-06-30
Support Year
14
Fiscal Year
2012
Total Cost
$270,709
Indirect Cost
$92,611
Name
Medical College of Wisconsin
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
937639060
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
Hudson, Stephen W; McNally, Mark T (2011) Juxtaposition of two distant, serine-arginine-rich protein-binding elements is required for optimal polyadenylation in Rous sarcoma virus. J Virol 85:11351-60
Van Dusen, Courtney M; Yee, Lily; McNally, Lisa M et al. (2010) A glycine-rich domain of hnRNP H/F promotes nucleocytoplasmic shuttling and nuclear import through an interaction with transportin 1. Mol Cell Biol 30:2552-62
Maciolek, Nicole L; McNally, Mark T (2008) Characterization of Rous sarcoma virus polyadenylation site use in vitro. Virology 374:468-76
McNally, Mark T (2008) RNA processing control in avian retroviruses. Front Biosci 13:3869-83
Maciolek, Nicole L; McNally, Mark T (2007) Serine/arginine-rich proteins contribute to negative regulator of splicing element-stimulated polyadenylation in rous sarcoma virus. J Virol 81:11208-17
Sun, Hai-Yuan; McNally, Mark T; Jackson, Vaughn E et al. (2006) Urea-nuclease treatment of concentrated retrovirions preserves viral RNA and removes polymerase chain reaction-amplifiable cellular RNA and DNA. J Virol Methods 137:304-8
McNally, Lisa M; Yee, Lily; McNally, Mark T (2006) Heterogeneous nuclear ribonucleoprotein H is required for optimal U11 small nuclear ribonucleoprotein binding to a retroviral RNA-processing control element: implications for U12-dependent RNA splicing. J Biol Chem 281:2478-88
Cochrane, Alan W; McNally, Mark T; Mouland, Andrew J (2006) The retrovirus RNA trafficking granule: from birth to maturity. Retrovirology 3:18
Maciolek, Nicole L; Alward, Wallace L M; Murray, Jeffrey C et al. (2006) Analysis of RNA splicing defects in PITX2 mutants supports a gene dosage model of Axenfeld-Rieger syndrome. BMC Med Genet 7:59
McNally, Lisa M; Yee, Lily; McNally, Mark T (2004) Two regions promote U11 small nuclear ribonucleoprotein particle binding to a retroviral splicing inhibitor element (negative regulator of splicing). J Biol Chem 279:38201-8

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