beta-Gal enzyme activity driven by the HSV LAT promoter does not correspond to beta-gal RNA levels in mouse trigeminal ganglia.
beta-Galactosidase enzyme expression can be detected in only a small percentage of trigeminal ganglia (TG) neurons acutely and latently infected with herpes simplex virus (HSV), in which the lacZ reporter gene was placed down-stream of the latency associated transcript (LAT) promoter at the LAT locus. However, DNA quantification suggests that a larger percentage of cells is infected than is expressing beta-galactosidase enzyme. To investigate the mechanism involved in regulation of genes expressed from the LAT promoter in trigeminal ganglia, in situ hybridization and histochemical staining assays were employed to determine on a cell-by-cell basis beta-gal gene expression both at the RNA and protein level. Using a LAT promoter-driven beta-gal construct in HSV-1 strain HFEM, it was found that there were 89-fold more cells positive for beta-gal transcript than cells positive for beta-gal enzyme in acutely infected trigeminal ganglia and a 10-fold difference in latently infected trigeminal ganglia. Thus, there is a discordance between beta-gal mRNA and beta-gal enzyme levels in HFEM/LAT-lacZ infected cells during acute and latent infection, and the beta-gal reporter gene activity does not faithfully compare the LAT promoter activity between acute and latently infected tissue. In contrast, in situ hybridization and histochemical staining assays were performed in mice acutely infected with a virus in which 140 bp of the LAT promoter sequences flanking the TATA element were replaced by 1.8 kbp of the neurofilament promoter (HSV-1 HFEM/NF-lacZ). This construct showed a correlation between beta-gal mRNA and enzyme expression in trigeminal ganglia in acute and latent infections. These findings suggest that sequences at the 5' end of the beta-gal transcript influence translation of the beta-gal message.
Huang
Q S
QS
Wistar Institute, Philadelphia, PA 19104, USA.
Valyi-Nagy
T
T
Kesari
S
S
Fraser
N W
NW
D000818:Animals; D001483:Base Sequence; D015870:Gene Expression; D005822:Genetic Vectors; D006651:Histocytochemistry; D017403:In Situ Hybridization; D007763:Lac Operon; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012313:RNA / Q000032:analysis*; D018139:Simplexvirus / Q000235:genetics*; D012668:Trigeminal Ganglion / Q000201:enzymology*; D001616:beta-Galactosidase / Q000235:genetics* / Q000378:metabolism*
D012313:RNA; D001616:beta-Galactosidase