Studies on the interactions mediated by the C-terminal zinc finger domains of Ikaros-family proteins

Abstract

Ikaros-family transcription factors play important roles in the control of haematopoiesis. Family members are predicted to contain up to six classical zinc fingers that are arranged into N- and C-terminal domains. The N-terminal domain is responsible for site-specific DNA binding, whereas the C-terminal domain mediates primarily the homo- and hetero-oligomerisation between family members. Although the mechanisms of action of these proteins are not completely understood, the zinc finger domains are known to play a central role. The work presented here was performed to further elucidate the physical and functional properties of the C-terminal domain, in particular, the C-terminal domain of Eos.

A range of biophysical techniques was used to demonstrate that the C-terminal domain of Eos mediates the formation of complexes that consist of nine or ten molecules. This result is highly unusual in the context of what is currently known about transcription-factor biology. In order to gain insight into the structural basis of oligomerisation, NMR spectroscopy was used to determine the solution structure of the second zinc finger from the C-terminal domain of Eos. This structure exhibited several unusual properties for a zinc finger, and these may contribute to the basis for the inability of this domain to bind DNA. Finally, alanine-scanning mutagenesis revealed residues that are involved in the homo-oligomerisation of E03, and these results are discussed in the context of the structure of the C-terminal domain. This work has not only shed light onto the mechanisms through which lkaros-family proteins regulate