Site-specific binding of proteins to DNA plays an important role in cell development, cell signaling, the cell cycle, and diseases such as cancer. There are two overarching goals to this proposal: (1) to develop a means for the identification of proteins bound to specific DNA sequences, and (2) to develop a better method for the identification of immunoprecipitation (IP) functional affinity reagents against DNA-bound proteins found on a complex antigen source. We propose to apply the proof-of-principle using the interaction of transcription factors (TFs) and their specific DNA binding sites as a model. TFs are one major example of such DNA-binding proteins. Once bound to their binding site(s), TF proteins can regulate the transcription of genes, thereby making individual genes either more or less active. Although this is a proposal to develop a better means toward obtaining functional affinity reagents against proteins when they are bound to DNA, the method itself can be applied to other uses.
Public Health Relevance Statement: Public Health Relevance: Many DNA-binding proteins are bound to specific DNA sequences and assist to regulate gene expression. Whether activating or repressing gene regulation, these proteins control the transfer of genetic information from DNA to RNA. As one example, transcription factor modulation of target genes leads to new molecules appearing on an immune cell's surface, including receptors for cytokines that stimulate cell proliferation and differentiation. We believe the proposed functional affinity screening method will allow us to more efficiently isolate affinity reagents that can be used to study the complex controlling mechanisms of the genome.
Project Terms: Affinity; Antibodies; Antigens; Binding (Molecular Function); Binding Proteins; Binding Sites; Biological Models; Cell Cycle; Cell Differentiation process; Cell Proliferation; Cell surface; Cells; Chip seq; Chromatin; Complex; Cytokine Receptors; design; Development; Disease; DNA; DNA Binding; DNA Sequence; DNA-Binding Proteins; Estrogen Receptors; Estrogens; Fibrinogen; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Transcription; Genome; Genomic DNA; Goals; Human; Immune; Immunoprecipitation; Individual; intercellular communication; Malignant Neoplasms; Mass Spectrum Analysis; Methods; Modeling; Nuclear; Play; Promotor (Genetics); Protein Binding; protein complex; Proteins; public health relevance; Reagent; research study; RNA; Role; screening; Site; Source; Test Result; Testing; transcription factor; Validation
