Two Leaky-Late HSV-1 Promoters Differ Significantly in Structural Architecture

Abstract

The HSV-1 VP5 and VP16 transcripts are expressed with leaky-late (gamma1) kinetics and reach maximal levels after viral DNA replication. While the minimal VP5 promoter includes only an Sp1 site at -48, a TATA box at -30, and an initiator (Inr) element at the cap site, here we show that elements upstream of -48 can functionally compensate for the mutational loss of the critical Sp1 site at -48. To determine whether this is a general feature of leaky-late promoters, we have carried out a detailed analysis of the VP16 promoter in the context of the viral genome at the gC locus. Sequence analysis suggests a great deal of similarity between the two. Despite this, however, mutational analysis revealed that the 5' boundary of the VP16 promoter extends to ca. -90. This region includes an Sp1 binding site at -46, CAAT box homology at -77, and "E box" (CACGTG) at -85. Mutational and deletional analyses demonstrate that the proximal Sp1 site plays little or no role in promoter strength; despite this it can be shown to bind Sp1 protein using DNA mobility shift assays. Like the VP5 promoter, the VP16 promoter also requires an initiator element at the cap site. The VP16 Inr element differs in sequence from that of the VP5 promoter, and its deletion or mutation has a significantly smaller effect on promoter strength. The difference between these two Inr elements was confirmed by our finding that the VP16 initiator element binds to the 65-kDa YY1 transcription factor, and the VP5 Inr element competes poorly for the binding between the VP16 element and infected cell proteins in comparative bandshift assays. While the VP16 Inr sequence is identical to that of several murine TATA-less promoters, the VP16 Inr requires a TATA box for measurable activity.

Analysis of Herpes Simplex Virus ICP0 Promoter Function in Sensory Neurons during Acute Infection, Establishment of Latency, and Reactivation In Vivo

“…Other null mutants at the gC and gJ loci have been shown to replicate normally in cell cultures (2,39,61). Therefore, any generalized replication defects in our mutants may indicate the presence of unknown secondsite mutations that could conceivably affect ICP0 promoter function.…”

“…However, they must lack sequences that are very important for expression in neurons but not in certain other cell types. Likely candidates include the initiator-like sequences near the mRNA start sites of leaky late promoters previously described (38,39) as well as downstream elements that have been shown to be important for the full expression of this class of HSV genes (3,4,72). Alternately, these promoters may contain neural tissue-specific down-regulatory signals that can be overcome by upstream sequences.…”

Transcription of the Herpes Simplex Virus 1 Genome during Productive and Quiescent Infection of Neuronal and Nonneuronal Cells

“…4E and F. This group of genes is more uniform in their expression relative to the other groups of genes and is represented by a minimum of 1% of the total reads (virus plus cell). The early and sustained high level of expression is most likely due to the presence of a TATA box INR and binding sites for upstream factors in their promoters (60). The binding sites for upstream factors enable expression early after infection, and the INR allows for the continued expression late after infection.…”

A Forward Phenotypically Driven Unbiased Genetic Analysis of Host Genes That Moderate Herpes Simplex Virus Virulence and Stromal Keratitis in Mice

“…A mechanism of molecular mimicry of an IgH locus and viral antigen leading to HSK has been described [47], but is limited to specific viral and mouse strains and dominant only under conditions of low viral load [53], which were not duplicated in our studies. Other potentially interesting genes within this region include YinYang1 (YY1), a transcription factor that can both up or down regulate genes through its cognate binding sites and is known to regulate HSV-1 genes [54], [55]. The YY1 gene contains a non-synonymous SNP between B6 and D2 and also has a cis expression QTL (cis-eQTL) at 110.06 Mb (determined with the Hippocampus Consortium data set in GN [56]), which regulates levels of expression in the nervous system.…”