Presentation description

DNA methylation is an abundant and reversible chemical modification of DNA that cells utilize to regulate gene transcription at CpG sites (mCpG), and it is essential for healthy cellular function. Alterations in DNA methylation patterns can lead to changes in gene expression and have been linked to various diseases, including several types of cancer. At many promoters, this epigenetic mark is often necessary but not sufficient on its own to alter gene expression. Methyl-CpG binding proteins (MBPs) recognize and bind to mCpG sites, recruiting additional proteins that regulate transcription. The three members of the ZBTB zinc finger (ZF) family of MBPs share a highly similar protein sequence in their ZF domain, but exhibit different preferences in the DNA sequences they recognize. ZBTB4 and ZBTB33 have demonstrated bimodal DNA recognition and selectively bind both TpG and mCpG DNA sequences through a conserved set of three ZFs. Despite the conservation, ZBTB4 demonstrates enhanced mCpG and TpG sequence selectivity, compared to ZBTB33. The mechanisms for DNA recognition have been structurally characterized for two ZF MBP family members, including ZBTB33 and ZBTB38. However, the molecular mechanism by which ZBTB4 recognizes its target DNA sequences remains uncharacterized. Herein, we present insights into ZBTB4 nucleic acid recognition and the potential role of non-conserved protein regions on DNA recognition. To do this, binding affinities of various ZBTB4 protein constructs were determined and compared using fEMSA (fluorescence-based electrophoretic mobility shift assays). These findings provide insight into how ZBTB4 selectively recognizes its genomic targets and expands upon ZF-MBP mechanisms of DNA recognition.