TOP2b directs and cooperates with XPG endonuclease to regulate t-RA inducible genes transcription

Transcription activation is a multistep process including spatial chromatin remodelling. We demonstrated that upon t-RA addition, RARA targets its RARE responsive element, to recruit TOP2B, while XPG/XPF are recruited sequentially after the general transcription factors and Pol II at the promoter of the activated gene. We found that TOP2B first generates DNA breaks in the RARE surroundings, a prerequisite for XPG to form a second set of DNA breaks at the promoter. ChIP and micro3C experiments further show that such DNA breaks, identified by Bio-UTP incorporation and gammaH2AX markers relax the DNA, allowing chromatin reorganisation to form some looping structures between RARE and promoter on the one side and promoter and gene terminator on the other side. These structures seems further stabilized by Mediator. Supporting this, silencing TOP2B almost completely abolishes the formation of DNA breaks at both RARE and promoter surroundings, while silencing XPG/XPF only prevent formation of DNA breaks at the promoter, ultimately collapsing RNA synthesis in both cases. This work highlights out the coordinated action and the leading role of TOP2B, together with XPG/XPF, in forming the transcription activation complex for the accurate expression of inducible genes.