Rotavirus is the major cause of severe, life-threatening gastroenteritis in young children and animals. Rotaviruses are large (1000 A), complex, icosahedral assemblies. This virus has been the subject of extensive biochemical, genetic and structural studies because of its medical relevance, intriguing structural complexity, and unique strategies of replication and morphogenesis. Rotaviruses contain 11 segments of double-stranded RNA encapsidated within three concentric capsid layers. Of the 12 proteins encoded by the genome, six are structural (VP1-7), and six are non-structural (NSP1-6). In the last three and half years, we have made exciting new discoveries that provide a better characterization of rotavirus structure and a deeper insight into the structural basis of various virus functions such as trypsin-enhanced infectivity, virus assembly, endogenous transcription, and genome replication and packaging. These recent developments, have allowed us to plan more in-depth dissection of structure-function correlations in rotavirus using a combination of highresolution cryo-EM, X-ray crystallographic, and biochemical techniques. The specific objectives of this ongoing project are: 1) To further investigate the mechanism of protease-enhanced infectivity and spike assembly, and conformational changes associated with cell entry of rotavirus using in vitro recombinant VLPbased techniques and electron cryo-tomographic approaches. 2) To further our understanding of the structural basis of endogenous transcription in rotavirus by characterizing the structural alterations in response to transcriptional activation using high-resolution cryo-EM techniques. 3) To dissect the structural mechanisms of regulation of rotavirus genome replication/packaging using X-ray crystallographic and single particle cryo-EM analysis of NSP2 with its various ligands such as RNA, NSP5, and VP1. 4) To explore further the role of the recently discovered novel NDP kinase activity in genome replication and to carry out structure-based design of inhibitors of this activity. 5) To construct an in vitro replication complex based on our hypothesis that capsid assembly and genome encapsidation in rotavirus are concomitant processes initiated by pentamers of dimers of VP2. 6) To carry out structural studies on rotavirus NSP1, a rotavirus non-structural protein that interferes with the interferon response pathway. Our structural information, in conjunction with continued advances in molecular virology of rotavirus, have the potential to enhance the development of more effective methods of disease prevention and control. More importantly, we expect to continue to discover new fundamental structural information to help understand how these complex viruses gain entry into host cells, assemble, transcribe, replicate, and package their genomes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37AI036040-24
Application #
8214561
Study Section
Special Emphasis Panel (NSS)
Program Officer
Cassels, Frederick J
Project Start
1988-12-01
Project End
2013-01-31
Budget Start
2012-02-01
Budget End
2013-01-31
Support Year
24
Fiscal Year
2012
Total Cost
$513,278
Indirect Cost
$178,895
Name
Baylor College of Medicine
Department
Biochemistry
Type
Schools of Medicine
DUNS #
051113330
City
Houston
State
TX
Country
United States
Zip Code
77030
Venkataram Prasad, B V; Air, Gillian M (2014) Editorial overview: virus-glycan interactions and pathogenesis. Curr Opin Virol 7:v-vi
Yu, Ying; Lasanajak, Yi; Song, Xuezheng et al. (2014) Human milk contains novel glycans that are potential decoy receptors for neonatal rotaviruses. Mol Cell Proteomics 13:2944-60
Ashline, David J; Yu, Ying; Lasanajak, Yi et al. (2014) Structural characterization by multistage mass spectrometry (MSn) of human milk glycans recognized by human rotaviruses. Mol Cell Proteomics 13:2961-74
Sastri, Narayan P; Viskovska, Maria; Hyser, Joseph M et al. (2014) Structural plasticity of the coiled-coil domain of rotavirus NSP4. J Virol 88:13602-12
Viskovska, Maria; Anish, Ramakrishnan; Hu, Liya et al. (2014) Probing the sites of interactions of rotaviral proteins involved in replication. J Virol 88:12866-81
Venkataram Prasad, B V; Shanker, Sreejesh; Hu, Liya et al. (2014) Structural basis of glycan interaction in gastroenteric viral pathogens. Curr Opin Virol 7:119-27
Criglar, Jeanette M; Hu, Liya; Crawford, Sue E et al. (2014) A novel form of rotavirus NSP2 and phosphorylation-dependent NSP2-NSP5 interactions are associated with viroplasm assembly. J Virol 88:786-98
Prasad, B V Venkataram; Schmid, Michael F (2012) Principles of virus structural organization. Adv Exp Med Biol 726:17-47
Baker, Matthew; Prasad, B V Venkataram (2010) Rotavirus cell entry. Curr Top Microbiol Immunol 343:121-48
Li, Zongli; Baker, Matthew L; Jiang, Wen et al. (2009) Rotavirus architecture at subnanometer resolution. J Virol 83:1754-66

Showing the most recent 10 out of 42 publications