![]() ![]() The former can be done using comprehensive, unbiased experimental and computational methods the latter requires more focused mechanistic analyses. ![]() To understand gene regulation and regulatory networks, it is therefore essential not only to accurately quantify these differences in DNA recognition but also to determine the structural and physical basis of that specificity. Nonetheless, subtle quantitative differences in DNA-binding specificity between related TFs are associated with large qualitative differences in the sets of target genes they control ( Maerkl and Quake 2007). 2014), leading to similar DNA sequence preferences among family members. The DNA-binding domains (DBDs) of TFs fall into families consisting of dozens or even hundreds of members ( Weirauch et al. Gene expression programs are precisely regulated by transcription factors (TFs), a class of DNA-binding proteins that orchestrate the activity of the RNA polymerase II and chromatin-modifying complexes. The binding models help explain differences in AR and GR genomic binding and provide a biophysical rationale for how promiscuous binding by GR allows functional substitution for AR in some castration-resistant prostate cancers. We conclude that the increased specificity of AR is correlated with more enthalpy-driven binding than GR. The specificity of AR extends to the regions flanking the core 15-bp site, where isothermal calorimetry measurements reveal that affinity is augmented by enthalpy-driven readout of poly(A) sequences associated with narrowed minor groove width. The relative preference between the two factors can be tuned over a wide range by changing the DNA sequence, with AR more sensitive to sequence changes than GR. This analysis revealed that the DNA-binding preferences of AR and GR homodimers differ significantly, both within and outside the 15-bp core binding site. We developed an algorithm, SelexGLM, that quantifies binding specificity over a large (31-bp) binding site by iteratively fitting a feature-based generalized linear model to SELEX probe counts. To address this paradox, we determined the intrinsic specificities of the AR and GR DNA-binding domains using a refined version of SELEX-seq. The DNA-binding interfaces of the androgen (AR) and glucocorticoid (GR) receptors are virtually identical, yet these transcription factors share only about a third of their genomic binding sites and regulate similarly distinct sets of target genes. ![]()
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