(From the Applicant?s Abstract): Recurrent herpes simplex virus type 1 (HSV-1) latent infection is a leading cause of viral induced corneal blindness. The mechanism of the HSV-1 latency-reactivation cycle remains elusive. The working hypothesis for this study is HSV-1 LAT?s anti-apoptosis activity plays a key role in LAT?s ability to enhance reactivation. To understand how LAT RNA protects cells against apoptosis and whether this LAT function correlates with its ability to enhance reactivation, the precise region of LAT that blocks apoptosis needs to be determined first. To fine map this region, a series of plasmids expressing different portion of the LAT RNA will be constructed and examined for anti-apoptosis activity by transient transfection assays. Some plasmids will be constructed in which the ATG of putative LAT potential open reading frames (ORFs) within the investigate LAT region will be altered by site directed mutagenesis to prevent translation of any potential ORF. Based on the above mapping experiments, a number of LAT mutants each expressing a key anti-apoptosis sequence (in an otherwise Lat negative virus) will be constructed to fine map the region of LAT responsible for anti-apoptosis activity in the context of the virus. Neuronal derived tissue culture cells will be infected and the ability of the Lat sequences to block HSV-1 induced apotosis will be assessed.
| Jin, Ling; Perng, Guey-Chuen; Brick, David J et al. (2004) Methods for detecting the HSV-1 LAT anti-apoptosis activity in virus infected tissue culture cells. J Virol Methods 118:9-13 |
