Identification and characterization of Hyphantria cunea nucleopolyhedrovirus homologous repeated regions

A total of six homologous regions (hycu-hrs1-6) were identified in the genome of Hyphantria cunea nucleopolyhedrovirus (HycuNPV). These hycu-hrs were localized in non-coding regions interspersed throughout the HycuNPV genome and showed structural homology to several other baculovirus hrs. Sequence analyses indicated that hycu-hrs were composed of 65-69-bp direct repeats, each of which contained a 29-31-bp imperfect palindrome embedded within non-palindromes tandemly arranged. hycu-hrs consisted of a variable number of repeats ranging from two for hycu-hr3 to twenty-five for hycu-hr6, and these repeats showed a high degree of identity to each other. The hycu-hr6, which was localized immediately upstream of HycuNPV gp64 gene (hycu-gp64), represented the longest hr among group I NPV hrs reported to date. Transient expression assays demonstrated that the expression of hycu-gp64 promoter-driven luciferase gene (luc) was dramatically enhanced by hycu-hr6 which was placed upstream and downstream of luc in an orientation-independent manner. Moreover, hycu-hr6-dependent enhancement was observed in the absence of any additional viral gene products, although it could be further strengthened in the presence of HycuNPV ie-1 gene product. These results indicate that hycu-hrs function as enhancers of transcription mediated by RNA polymerase II, and suggest for the first time that efficiency of gp64 promoter is dependent on the enhancement function of hrs.