Scott Shors, PhD Department of Biology New Mexico Institute of Mining and Technology sshors@nmt.edu Phone:(505) 835-6187 Personal Website

Title: Role of Stress Granule Formation in Mediating Viral Infection

In response to viral infection, the host attempts to limit virus production by inhibiting protein synthesis.  At least two kinases can limit viral infection; PKR and Perk.  Both of these kinases inhibit protein synthesis by phosphorylating the translation initiation factor eIF2.  As the amount of eIF2 that is phosphorylated increases and is unable to participate in protein synthesis, not only does the total amount of protein translated decrease, but the profile of what proteins are translated changes.  A key aspect of this profile change is the result of the action of two proteins; TIA-1 and TIA-R.  When the amount of eIF2 available for translation drops to a threshold level, these two proteins sort the mRNA that encoded the so-called 'housekeeping' genes to cytoplasmic bodies termed stress granules (SG).  TIA-1 and TIA-R do not sequester the stress response transcripts.  This aspect of the innate immune response will limit viral production unless the virus has some way of circumventing the SG response.  Very little is known about the role of SG formation in controlling viral infection.  By using TIA-1 or TIA-R knockout cell lines we have preliminary data that TIA-R is not involved in the innate response against herpes simplex 1 (HSV1) whereas TIA-1 plays a critical role in limiting HSV1 production (i.e. viral titers for HSV1 are similar in both wild type cells and TIA-R-/- cells, but significantly higher in TIA-1-/- cells).  Furthermore, we have data that demonstrates that the ability of TIA-1 to limit HSV1 infection is strain dependant.  Both the SP7 strain, which is neuroinvasive, and the attenuated KOS strain lead to similar titers in wild type cells, but in TIA-1 knockout cells, SP7 titers increase by over 900 fold while KOS titers only increase 40 fold.  In contrast to HSV1, vaccinia virus is inhibited by TIA-R and not by TIA-1.

Specific Aims:

Aim 1:  Does TIA-1 or TIA-R play a role in inhibiting viral replication under single cycle growth conditions?  In addition to testing other strains of HSV1 and vaccinia virus the following viruses will also be tested: adenovirus, reovirus, adeno-associated virus, picornavirus, rhabdovirus, arenavirus, Influenza A, and retrovirus.

Aim 2:  Does TIA-1 or TIA-R play a role in inhibiting viral replication under single cycle growth conditions in cells that have been pretreated with 100 units of interferon alpha?  The same viruses used in Aim 1 will be tested.

Aim3 / Future studies:  Determining what viral proteins and transcripts interact with TIA-1 and TIA-R via SDS-PAGE, western blotting, northern blots and PCR.  This data will shed light on a basic part of the innate immune response and aid in design of vaccine strains and gene therapy vectors.