β-Gal enzyme activity driven by the HSV LAT promoter does not correspond to β-gal RNA levels in mouse trigeminal ganglia

Abstract

β-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 downstream 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 β-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 β-gal gene expression both at the RNA and protein level. Using a LAT promoter-driven β-gal construct in HSV-1 strain HFEM, it was found that there were 89-fold more cells positive for β-gal transcript than cells positive for β-gal enzyme in acutely infected trigeminal ganglia and a 10-fold difference in latently infected trigeminal ganglia. Thus, there is a discordance between β-gal mRNA and β-gal enzyme levels in HFEM/LAT-lacZ infected cells during acute and latent infection, and the β-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 β-gal mRNA and enzyme expression in trigeminal ganglia in acute and latent infections. These findings suggest that sequences at the 5′ end of the β-gal transcript influence translation of the β-gal message.