|De Los Santos, Teresa|
|Diaz-san Segundo, Fayna|
Submitted to: Journal of Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/24/2008
Publication Date: 12/3/2008
Citation: De Los Santos, T.B., Diaz-San Segundo, F., Zhu, J.J., Koster, M.J., Dias, C., Grubman, M.J. 2008. A Conserved Domain in the Leader Proteinase of Foot-and-Mouth Disease Virus is Required for Proper Sub-Cellular Localization and Function. Journal of Virology. 83(4):1800-1810. Interpretive Summary: Foot-and-Mouth Disease Virus (FMDV) causes an economically devastating disease of cloven-hoofed animals. To develop strategies necessary to control this disease it is important to understand how the virus and host interact. Identification of mechanisms that the host can utilize to rapidly control and contain virus replication may result in disease control approaches that are able to augment current and potential new vaccine strategies. We have developed a weakened version of FMDV (leaderless virus) that lacks a portion of the viral genome (lacks the leader protein coding region). This virus is highly attenuated when inoculated into cattle or swine, but can replicate to a limited extent. However, after aerosol infection of cattle the leaderless virus remained localized in the lungs and in contrast to virulent virus did not spread to other areas of the animal and cause disease. In certain cell cultures, we found that leaderless virus infection induced a host antiviral response that blocked virus spread and we have identified host proteins that directly inhibit virus replication. Over the past few years we have initiated a program to understand the mechanism by which leaderless virus is attenuated. We have demonstrated that the FMDV leader protein is involved in degrading a cellular protein, nuclear factor-kappa B (NF-'B), which participates in inducing the host cell antiviral response. Degradation of this host protein occurs in the nucleus. Thus, the presence of FMDV L protein and its movement and maintenance in the nucleus allows the virus to overcome the protective host response and replicate to high levels. To gain a more comprehensive understanding of the mechanism of L protein inhibition of the host antiviral response we used a bioinformatics approach and made mutations in a potential motif of L protein that is involved in its maintenance in the nucleus. The mutated virus does not degrade NF-'B and is attenuated in tissue culture. Understanding the mechanism of FMDV inhibition of the host antiviral response at the molecular level should be helpful in the development of specific antiviral strategies that can rapidly inhibit or limit virus spread.
Technical Abstract: The leader proteinase (Lpro) of foot-and-mouth disease virus (FMDV) is involved in antagonizing the innate immune response by blocking the expression of interferon (IFN) protein and by reducing the immediate-early induction of IFN beta mRNA and IFN stimulated genes. In addition to its role in shutting off cap dependent host mRNA translation, Lpro is associated with the degradation of the p65/RelA subunit of the transcription factor NF-kappa B Bioinformatic analysis suggests that Lpro contains a SAP (SAF-A/B, Acinus and PIAS) domain, a protein structure associated in some cases, with the nuclear retention of molecules involved in transcriptional control. We have mutated conserved Lpro residues within this domain and obtained stable mutant viruses that displayed an attenuated phenotype. Indirect immuno-fluorescence analyses showed that Lpro sub-cellular distribution is altered in cells infected with a mutant virus. Interestingly, nuclear p65/RelA staining disappeared from wild type (WT) but not from mutant virus infected cells. Consistent with these results, NF-kappa B dependent transcription was not inhibited in cells infected with mutant virus in contrast to cells infected with WT virus. However, degradation of the translation initiation factor eIF-4G was very similar for the both, WT and mutant, viruses. Since Lpro catalytic activity was demonstrated to be a requirement for p65/RelA degradation, our results indicate that mutation of the SAP domain revealed a novel separation-of-function activity for FMDV Lpro.