Reverse Regulation: Controlling Intrinsically Disordered Domains with Structured Elements

Intrinsic disorder in proteins correlates with alternatively spliced motifs, protein interaction domains, and post-translational modification sites. Consequently, there are many examples of intrinsically disordered regions sensing multiple cellular signals and responding by modulating the activity of a structured functional domain. Conversely, we have discovered two examples of the reverse process - regulation of an intrinsically disordered domain by a structured protein element - using the Drosophila Hox transcription factor Ultrabithorax (Ubx) as a model system. Both in silico and in vitro approaches identified a large (∼150 a.a.) intrinsically disordered domain within the Ubx transcription activation domain, which is bounded on its C-terminus by an alpha helix. In cell culture promoter-reporter assays, point mutations that enhance helix stability increase transcription activation, whereas mutations that destroy helix structure abrogate transcription activation, leaving repression and DNA binding intact. Indeed, two amino acid changes are sufficient to disable a 150 a.a. intrinsically disordered domain. These mutations alter Ubx function in a tissue-dependent manner in Drosophila, emphasizing the fact that in silico prediction and in vitro characterization of intrinsically disordered domains is relevant to the function of the protein in a live animal. In the second example, we monitored DNA binding as a function of osmotic stress to discover DNA binding triggers a conformational change that exposes significant additional surface area in N-terminal half of Ubx, including the intrinsically disordered domain. This conformational change provides an opportunity for DNA, via the structured DNA-binding homeodomain, to impact both transcription regulation and protein interactions by the intrinsically disordered domain. This regulatory mode could potentially select the mode (activation vs. repression) of transcription regulation by Ubx in response to DNA sequence.