Effect of G-quadruplex loop mutations on the G-quadruplex formation, protein binding and transcription of BmPOUM2 in Bombyx mori

Abstract

A G-quadruplex (G4) was identified in the promoter of transcription factor BmPOUM2 in Bombyx mori. This G4 structure contains three loops and is bound by transcription factor BmLARK, facilitating the transcription of BmPOUM2. However, the relationship between the structure and function of the BmPOUM2 G4 remains to be clarified. In this study, loop mutants of the BmPOUM2 G4 structure were generated to study the function of the structure in transcription regulation. The results revealed that mutations of Loops A and B could not completely suppress G4 formation, but affected the binding of the G4 structure with BmLARK and the promoter activity. The mutation (C-to-T) of the one-nucleotide-loop, Loop C, enhanced the G4 formation, its binding with BmLARK and the transcription activity of the BmPOUM2 promoter. It is speculated that the binding site of BmLARK probably is on the G-quartet planes, rather than on the loops, which may assist the maintenance and modification of the G4 structure and its protein binding activity.

Keywords: G-quadruplex; loop; mutation; protein binding; silkworm; transcription.