Transcription factor Sp2 potentiates binding of the TALE homeoproteins Pbx1:Prep1 and the histone-fold domain protein Nf-y to composite genomic sites

Different transcription factors operate together at promoters and enhancers to regulate gene expression. Transcription factors either bind directly to their target DNA or are tethered to it by other proteins. The transcription factor Sp2 serves as a paradigm for indirect genomic binding. It does not require its DNA-binding domain for genomic DNA binding and occupies target promoters independently of whether they contain a cognate DNA-binding motif. Hence, Sp2 is strikingly different from its closely related paralogs Sp1 and Sp3, but how Sp2 recognizes its targets is unknown. Here, we sought to gain more detailed insights into the genomic targeting mechanism of Sp2. ChIP-exo sequencing in mouse embryonic fibroblasts revealed genomic binding of Sp2 to a composite motif where a recognition sequence for TALE homeoproteins and a recognition sequence for the trimeric histone-fold domain protein nuclear transcription factor Y (Nf-y) are separated by 11 bp. We identified a complex consisting of the TALE homeobox protein Prep1, its partner PBX homeobox 1 (Pbx1), and Nf-y as the major partners in Sp2-promoter interactions. We found that the Pbx1:Prep1 complex together with Nf-y recruits Sp2 to co-occupied regulatory elements. In turn, Sp2 potentiates binding of Pbx1:Prep1 and Nf-y. We also found that the Sp-box, a short sequence motif close to the Sp2 N terminus, is crucial for Sp2's cofactor function. Our findings reveal a mechanism by which the DNA binding-independent activity of Sp2 potentiates genomic loading of Pbx1:Prep1 and Nf-y to composite motifs present in many promoters of highly expressed genes.