The Chilo iridescent virus DNA polymerase promoter contains an essential AAAAT motif

The delayed-early DNA polymerase promoter of Chilo iridescent virus (CIV), officially known as Invertebrate iridescent virus, was fine mapped by constructing a series of increasing deletions and by introducing point mutations. The effects of these mutations were examined in a luciferase reporter gene system using Bombyx mori cells transfected with promoter constructs and infected with CIV. When the size of the upstream element was reduced from position ¿19 to ¿15, relative to the transcriptional start site, the luciferase activity was reduced to almost zero. Point mutations showed that each of the 5 nt (AAAAT) located between ¿19 and ¿15 were equally essential for promoter activity. Mutations at individual bases around the transcription initiation site showed that the promoter extended until position ¿2 upstream of the transcription start site. South-Western analysis showed that a protein of approximately 100 kDa interacted with the ¿19 nt promoter fragment in CIV-infected cells. This binding did not occur with a point mutant that lacked promoter activity. The AAAAT motif was also found in the DNA polymerase promoter region of other iridoviruses and in other putative CIV delayed-early genes