Study of molecular structural and dynamical aspects of transcriptional regulatory factors

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The goal of this project is to elucidate molecular mechanisms for regulating the enhanceosome formation, particularly, stabilization of transcription factors bound to an enhancer region of a gene.We investigated some complexes containing a transcriptional regulatory factor, Runx1, which plays a pivotal role in hematopoiesis and is involved in acute leukemias. The DNA binding activity of Runxl is enhanced allosterically by non-DNA binding heterodimeric partner, CBFβ. Previously, we showed that there was no significant structural change of Runxl upon binding of CBFβ when a crystal structure of Runx1-DNA complex was compared with that of Runx1-CBFβ-DNA complex. It promoted us to focus on molecular fluctuations of Runx1. Magnetic relaxation measurements using NMR demonstrated that some regions of Runx1 are significantly fluctuated without CBFβ, while the fluctuations of Runx1 became reduced upon binding of CBFβ. We proposed that regulations of molecular functions are accomplished by alteri … More ng molecular fluctuations.Runxl is also involved in transcriptional regulation of the T cell antigen receptor a chain (TCRα) together with another transcriptional regulatory factor, Etsl. The DNA binding activity of Etsl is controlled by an N-terminal flanking region of ETS domain, referred exon VII. The exon VII inhibits the DNA binding activity of Ets1 while the inhibition is released by an adjacently placed Runxl. Etsl is not capable of binding the enhancer upon phosphorylation of exon VII even if Runxl binds to the enhancer. We investigated the molecular details of the complex composed of Runxl and Etsl in the TCRα enhancer using X-ray crystallography, EMSA and the luciferase reporter gene assay. We identified which regions of Etsl are responsible for cooperative DNA binding with Runxl and the impact of the exon VII phosphorylation. Further structural analyses might be required to unveil overall aspect of the regulatory systems. We will also study about fluctuations of Runxl upon Etsl binding. Less