Structural studies in mammalian gene regulation

Abstract

Functional complexity in higher eukaryotes is thought to be attributable to increasing sophistication in gene expression patterns, giving rise to highly specialised cell types and tissues. Regulation of gene expression is achieved through diverse mechanisms. The research in this thesis addresses two aspects of gene regulation in mammals: the interactions of classical zinc finger proteins with RNA, and the mechanisms by which transcriptional regulators are recruited to specific chromatin sites. Firstly, the RNA-binding properties of the classical zinc finger transcription factor Yin-Yang 1 (YY1) are examined. Accumulating evidence suggests that interactions of transcription factors with RNA might play a role in gene regulation. Previous studies have shown that full-length YY1 can bind various RNA species, but little is known about the molecular detail of these interactions. The work in this thesis demonstrates that the classical zinc fingers of YY1 bind RNA, and surveys their RNA sequence preferences. It also begins to elucidate the mode of RNA recognition employed by YY1 zinc fingers, revealing an RNA-binding interface that is largely distinct from canonical DNA-binding residues. Secondly, the interaction between bromodomain-containing protein 3 (Brd3) and the chromatin remodelling enzyme CHD4 is explored. Brd3 bromodomains recognise acetylated lysines on histones, a property which has led to its characterisation as an epigenetic ‘reader’ that serves as the link between chromatin modifications and transcriptional outcomes. This function necessitates the interaction of Brd3 with other transcriptional effector proteins, but the mechanisms of effector recruitment by Brd3 are currently poorly understood. In the present work, the structure of a complex between Brd3 and CHD4 was determined, yielding insights into the molecular basis of Brd3-mediated recruitment of transcriptional regulators.