Establishment and maintenance of HSV latent infection is mediated through correct splicing of the LAT primary transcript.
To study the effect of the splicing of HSV-1 latency-associated transcript (LAT) on viral latency, we constructed two mutant viruses (FHlambda+ and FHlambda-) in which the 168-bp HpaI-HpaI fragment within the 2-kb LAT intron was replaced by a 447-bp bacteriophage lambda sequence. The lambda DNA was inserted in opposite orientations in FHlambda+ and FHlambda-. The mutation in FHlambda+ disrupted the splicing of LAT primary transcript and altered both LAT exon and intron, whereas the mutation in FHlambda- virus preserved the wild-type splice sites and the wild-type exon. Quantitative PCR analysis revealed that during latency there was a reduction in the number of viral genomes in mouse trigeminal ganglia infected with FHlambda+ but not in those infected with FHlambda-. The decrease in the latent genome numbers was not due to a defect in viral replication during the acute stage of infection. Furthermore, trigeminal ganglia from mice latently infected with FHlambda+ displayed a slower reactivation kinetics compared to those infected with the parental strain. To elucidate the mechanism, we examined the antiapoptotic properties of these LAT constructs. A plasmid containing the pHlambda+ construct was found to be less protective for cells against apoptosis than plasmid containing the wild-type or pHlambda- constuct. These results suggest that the splicing of LAT primary transcript, and thus the correctly spliced exon product, play an important role in promoting the establishment and/or maintenance of viral latency.
Kang
Wen
W
Department of Microbiology, University of Pennsylvania Medical School, 315 Johnson Pavilion, Philadelphia, PA 19104-6076, USA.
Mukerjee
Ruma
R
Fraser
Nigel W
NW
D000818:Animals; D002522:Chlorocebus aethiops; D015967:Gene Expression Regulation, Viral; D006561:Herpes Simplex / Q000821:virology; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000502:physiology*; D051379:Mice; D009154:Mutation / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics*; D014158:Transcription, Genetic / Q000235:genetics*; D012668:Trigeminal Ganglion / Q000821:virology; D014709:Vero Cells; D014775:Virus Activation; D017735:Virus Latency / Q000235:genetics* / Q000502:physiology*