This is an application for renewal of a grant to study picornavirus genome replication. Picornaviruses represent an existing and emerging threat to US public health. Although protein factors and genetic elements required for picornavirus genome replication are known and appear to be conserved, a clear understanding of the mechanisms employed to produce picornaviral RNA is lacking. The long-term objective of this program is to reconstitute picornavirus genome replication in vitro from purified components. We have achieved all of the major objectives of the previous funding period. In addition, we have solved the first crystal structure for a picornaviral 3CD protein, developed the technology to study 3C-RNA interactions by using nuclear magnetic resonance spectroscopy, and discovered that the 3CD protein has both pre- and post-replication functions. During the next funding period, we will continue our studies of picornavirus genome replication as well as explore our newly discovered function for 3CD by pursuing the following specific aims: (1) Define the mechanism of assembly and structural organization of the picornavirus VPg uridylylation complex by using molecular genetic, biochemical and biophysical approaches;(2) Define the molecular basis for sequence- and structure-specific RNA recognition by 3C by using nuclear magnetic resonance spectroscopy;and (3) Elucidate the function of 3CD in formation of replication complexes.

Public Health Relevance

Picornaviruses represent an existing and emerging threat to US public health. Achievement of the goals of the application will provide novel targets and mechanisms for development of inhibitors to treat infections by picornaviruses, especially those for which vaccines are not available.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI053531-08
Application #
8013008
Study Section
Special Emphasis Panel (ZRG1-GGG-A (90))
Program Officer
Park, Eun-Chung
Project Start
2003-07-01
Project End
2013-01-31
Budget Start
2011-02-01
Budget End
2012-01-31
Support Year
8
Fiscal Year
2011
Total Cost
$472,568
Indirect Cost
Name
Pennsylvania State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
003403953
City
University Park
State
PA
Country
United States
Zip Code
16802
Chan, Yan M; Moustafa, Ibrahim M; Arnold, Jamie J et al. (2016) Long-Range Communication between Different Functional Sites in the Picornaviral 3C Protein. Structure 24:509-17
Moustafa, Ibrahim M; Gohara, David W; Uchida, Akira et al. (2015) Conformational Ensemble of the Poliovirus 3CD Precursor Observed by MD Simulations and Confirmed by SAXS: A Strategy to Expand the Viral Proteome? Viruses 7:5962-86
Kolli, Swapna; Meng, Xiangzhi; Wu, Xiang et al. (2015) Structure-function analysis of vaccinia virus H7 protein reveals a novel phosphoinositide binding fold essential for poxvirus replication. J Virol 89:2209-19
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Cameron, Craig E; Oh, Hyung Suk; Moustafa, Ibrahim M (2010) Expanding knowledge of P3 proteins in the poliovirus lifecycle. Future Microbiol 5:867-81
Hsu, Nai-Yun; Ilnytska, Olha; Belov, Georgiy et al. (2010) Viral reorganization of the secretory pathway generates distinct organelles for RNA replication. Cell 141:799-811
Pathak, Harsh B; Oh, Hyung Suk; Goodfellow, Ian G et al. (2008) Picornavirus genome replication: roles of precursor proteins and rate-limiting steps in oriI-dependent VPg uridylylation. J Biol Chem 283:30677-88
Shen, Miaoqing; Reitman, Zachary J; Zhao, Yan et al. (2008) Picornavirus genome replication. Identification of the surface of the poliovirus (PV) 3C dimer that interacts with PV 3Dpol during VPg uridylylation and construction of a structural model for the PV 3C2-3Dpol complex. J Biol Chem 283:875-88

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