Eukaryotic transcription hinges on the conceptual framework of combinatorial control, whereby the transcriptional output depends on the coordinated activities of multiple sequence specific transcription factors. The arrangement of cis-regulatory elements, protein-DNA contacts and the interplay of different transcriptional regulators through protein-protein interactions cumulatively operate towards the assembly of nucleoprotein complexes. A comprehensive understanding of the molecular mechanisms underlying multiprotein complex formation is thus vital to grasp the principles transcription rests on. Animals develop using the same key transcriptional regulators, with the highly conserved family of Hox transcription factors enacting a universal role to sculpt animal body plans along their anterior-posterior axis. In this Dissertation, we define some of the molecular principles that sculpt Hox-containing complex assembly. We show that alternative splicing of homothorax ( hth) generates two types of protein isoforms, one that contains a DNA binding homeodomain (HthFL) and one that does not contain a homeodomain (HDless). Both types of Hth isoforms include the evolutionarily conserved HM domain, which mediates a direct interaction with Extradenticle (Exd), another homeodomain protein. We show that although both HthFL and HDless isoforms of Hth can induce the nuclear localization of Exd, they carry out distinct sets of functions during development. Thus, alternative splicing of hth results in the generation of multiple transcription factors that execute unique functions in vivo. We further demonstrate that the mouse ortholog of hth, Meis1, also encodes a HDless isoform, suggesting that homeodomain-less variants of this gene family are evolutionarily ancient. Furthermore, we present evidence that the activity of the Hox protein Abdominal-A (Abd-A) is dependent on sequences outside of the highly conserved HD and YPWM motif. Namely, the C-terminus serves a core role in both the instruction of epidermal fates, as well as the repression of the Abd-A target gene Distalless (Dll) in vivo. Our observations underscore the importance of regions outside the HD for Abd-A activity, arguing that sequences beyond the HD are not all neutral to protein function but play a decisive role in sculpting the overall properties of Hox as transcriptional regulators. We also demonstrate that Abd-A overrides the activity of more anterior Hox proteins, a phenomenon called phenotypic suppression, utilizing this novel C-terminal domain and that dominance rests on an Exd-dependent DNA binding mechanism.
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