Characterizing interactions between cAMP responsive element binding protein 1 and methyl-CpG-binding protein 2 as a potential transcriptional activation complex


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Transcriptional activity is controlled by many types of DNA binding proteins. In addition to transcription factors that activate transcription by recruitment of RNA polymerase II, there are proteins like methyl-CpG-binding protein 2 (MeCP2). MeCP2 regulates transcription by binding to methylated DNA. MeCP2 is traditionally associated with being a transcriptional repressor by binding to methylated CpG dinucleotides and recruiting corepressors. Literature has shown that MeCP2 is also a transcriptional activator and has been proposed to bind directly to cAMP responsive element binding protein (CREB1) to facilitate this action. This hypothesis is under-studied biochemically, and this project aims to elucidate biochemical and biophysical information to further understand this potential interaction. In this project, we qualitatively study protein-protein interactions between MeCP2 and CREB1 in solution through native polyacrylamide gel electrophoresis. We also examine this hypothesis quantitatively by examining the effects of MeCP2 on CREB1 binding to DNA through fluorescence anisotropy assays. These studies suggest that there is a potential interaction between MeCP2 and CREB1. In addition to examining the presence of interactions between MeCP2 and CREB1, we also make progress on establishing protocols for studying the effects of nucleosomes on transcription factor target search. In summary, we establish purification protocols for CREB1 and MeCP2, preparation protocols for nucleosome core particle reconstitution, and examine interactions between CREB1 and MeCP2 through quantitative and qualitative methods.



CREB1, MeCP2, transcriptional activation, protein-protein interactions, DNA binding proteins