Unravelling Mechanistic Insights of Eukaryotic Transcriptional Control

Abstract

DNA transcription involves the association of multiple proteins with DNA to convert the genetic information into messenger-RNA transcripts, which later will be translated into proteins. Con-trolled regulation of transcription is imperative for preserving the healthy and homeostatic state of cells. In eukaryotes, where the accessibility of DNA is limited by the chromatin packaging, transcriptional control occurs primarily at the initiation stage. The initiation of transcription is regulated by a class of proteins termed transcription factors, which recognise and bind with high specificity short non-coding DNA fragments, so-called response elements, to modulate the re-cruitment of the transcription machinery. The mapping of how transcription factors achieve binding specificity for their genomic target sites, shielded by the massive amount of non-specific DNA, remains incomplete despite decades of dedicated research. Over the years, traditional structure determination techniques have provided multiple insights into specificity of transcrip-tion factors-DNA interactions, though from a more static perspective. We still lack information on why transcription factors bind related sequences with different affinity and unrelated se-quences with similar affinity. Furthermore, the response elements occur more frequently in the genome than the actual genes regulated by the transcription factors. To understand these aspects of transcriptional control, it is imperative also to study the dynamic aspects of the transcription factors-DNA interactions. Until relatively recently, DNA has been looked at as a rather passive component in the recognition process. However, DNA is highly polymorphic and its flexibility is sequence specific, allowing DNA to adapt in response to various conditions. In addition, DNA state constantly changes during the life time of a cell as a consequence of DNA supercoiling, the predominant regulatory force of transcriptional control. DNA supercoiling, introduced by molec-ular motors, can propagate along the chromatin fibre, altering the accessibility of the genetic code, which in turn could affect the binding of transcription factors. Taken together, reveals DNA as the key driver of many biological processes. In this thesis, I use computational meth-ods, to unravel mechanistic details of eukaryotic transcriptional control. I study naked DNA and DNA interactions with various transcription factors, in particular BZIP- and BHTH-factors. In my quest, I have uncovered the mechanism of how DNA epigenetic modifications and transcrip-tion factors binding modulate the torsional rigidity of DNA; and how these properties can be utilised in the regulation of transcription. I have also contributed to the understanding of how transcription factors exploit the local sequence specific plasticity of DNA in recognition of their genomic target sites, and how small alterations in DNA flexibility due to DNA methylation could make their targets sites unrecognisable. The major conclusion from this thesis work, is that the enigma of selective binding of transcription factors has its solution in the dynamics of transcrip-tion factor-DNA complexation combined with sequence specific DNA flexibility of response elements but also their flanking sites. The flanking sites modulate the conformational adaptability of the response elements, making them more shape-complementary for the transcription factor to form specific contacts, which increases the stability of the complex. The level of conformational adaptability of the DNA response elements to the binding by transcription factor, could modulate the degree of torsional rigidity of DNA to undergo supercoiling transitions, which in turn may modulate the recruitment of different collaborative transcription factors control how long gene promoters stay open and consequently being transcribed.