The relationship of herpes simplex virus latency associated transcript expression to genome copy number: A quantitative study using laser capture microdissection
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To investigate the quantitative relationship of latent herpes simplex virus (HSV) genomes to the expression of latency associated transcripts (LATs) we used a combination of laser capture microdissection (LCM), polymerase chain reaction (PCR), and quantitative real-time PCR to determine the number of HSV genomes in individual neurons of the mouse trigeminal ganglion (TG) during viral latency. Both LAT-positive and LAT-negative neurons detected by in situ hybridization (ISH) and lifted by LCM contained HSV genomes detected by PCR for HSV ICP47. The number of genomes/cell determined by real-time PCR with probes for HSV UL44 following LCM demonstrated a Poisson distribution with a predicted mean count of 178 genomes /LAT-positive neuron, and 68 genomes/LAT-negative neuron. The range was similar between the LAT-positive and LAT-negative neurons, and there was a substantial overlap in the distributions. These results suggest that the expression of LATs in an amount that is detectable by ISH does not depend only on the number of HSV genomes in the cell, and by implication suggests that neuron-specific factors play a role in the regulation of LAT expression during latency.
Keywordsherpes simplex virus latency latency associated transcript (LAT) real-time PCR laser capture microdissection
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- Cohrs RJ, Randall J, Smith J, Gilden DH, Dabrowski C, van Der Keyl H, Tal-Singer R (2000). Analysis of individual human trigeminal ganglia for latent herpes simplex virus type 1 and varicella-zoster virus nucleic acids using realtime PCR. J Virol 74(24): 11464–11471.PubMedCentralPubMedCrossRefGoogle Scholar
- Hill JM, Gebhardt BM, Wen R, Bouterie AM, Thompson HW, O’Callaghan RJ, Haiford WP, Kaufman HE (1996). Quantitation of herpes simplex virus type 1 DNA and latency-associated transcripts in rabbit trigeminal ganglia demonstrates a stable reservoir of viral nucleic acids during latency. J Virol 70(5): 3137–3141.PubMedCentralPubMedGoogle Scholar
- Roizman B, Sears A (1996). Herpes simplex viruses and their replication. In: Fields Virology. Fields BN, Knipe DM, Howley PM, Chanock RM, Hirsch MS, Melnick JL, Monath TP, Roizman B (eds). 3rd. ed., vol. 2, Philadelphia: Lippincott-Raven, pp 2231–2295.Google Scholar
- Trousdale MD, Steiner I, Spivack JG, Deshmane SL, Brown SM, MacLean AR, Subak-Sharpe JH, Fraser NW (1991). In vivo and in vitro reactivation impairment of a herpes simplex virus type 1 latency-associated trancript variant in a rabbit eye model. J Virol 65: 6989–6993.PubMedCentralPubMedGoogle Scholar