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United States Department of Agriculture

Agricultural Research Service

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Location: Foreign Animal Disease Research

2013 Annual Report

1a. Objectives (from AD-416):
The objective of this collaborative research project is to identify the host proteins that play critical roles in the process of virus replication. Identified host proteins will be characterized in terms of cell location, interaction’s kinetics, and participation of the virus cycle. Additionally, the mapping of the specific areas of the host proteins responsible of the interaction with the virus proteins will be identified.

1b. Approach (from AD-416):
The identified host proteins will be identified by the two hybrid yeast system. Those identified will be further characterized, particularly the molecular interaction between the host and virus proteins. Residues in the host proteins mediating the interaction with the virus proteins will be identified and characterized. Based on that information, the role of those residues in the process of virus replication will be assessed. Monoclonal antibodies specific for the different virus proteins are critical tools for this type of study since they allow the identification of the virus proteins in different methodological procedures to study the characteristics of the interaction between the host and virus proteins.

3. Progress Report:
The purpose of this collaborative research agreement between ARS PIADC and ISZLER is to better understand the role of Foot-and-Mouth Disease Virus (FMDV) proteins in the process of virus replication and pathogenesis in the natural host. IZLER provided ARS PIADC with a complete set of monoclonal antibodies (mAb) well characterized in terms of their specificity and reactivity to FMDV structural and non-structural proteins. These mAb were used with a yeast two-hybrid methodology to identify host proteins that interact with viral proteins. Several host proteins were identified which interacted with some of the virus proteins. In the particular case of FMDV non structural protein 2C we identified for first time, two different host proteins Beclin 1 and Vimentin, which were shown to be critical in the process of virus replication. Interactions of 2C with both host proteins were further characterized in FMDV-infected cells. Alanine scanning mutagenesis was used to map the specific amino acid residues in 2C critical for Beclin1 and Vimentin binding. Using reverse genetics, we identified 2C residues that in both cases are necessary for virus growth, suggesting that the interaction between FMDV 2C and cellular Beclin1 and Vimentin is essential for virus replication. No technologies were transferred in FY 2013. Two manuscripts containing these results have been published: Foot and Mouth Disease Virus non structural protein 2C interacts with Beclin1 modulating virus replication. D.P. Gladue, V. O’Donnell, R. Baker-Branstetter, L.G. Holinka, J. Pacheco, I. Fernandez-Sainz, Z. Lu, E. Brocchi, B. Baxt, M.E. Piccone, L. Rodriguez, and M.V. Borca. Journal of Virology (2012) 86:12080-12090 Foot-and-mouth disease virus modulates cellular vimentin for virus survival. D. P. Gladue, V. O’Donnell, R. Baker-Branstetter, L. G. Holinka, J. M. Pacheco-Tobin, I. Fernandez Sainz, Z. Lu, L. Rodriguez, and M. V. Borca. Journal of Virology (2013) 87:6794-803. During FY 2014 futher assessement of the structural and non-structual proteins will be conducted utilzing the mAbs provided by IZLER.

4. Accomplishments

Last Modified: 05/27/2017
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