Doublesex and the regulation of sexual dimorphism in Drosophila melanogaster: structure, function, and mutagenesis of a female-specific domain.
The DSX (Doublesex) transcription factor regulates somatic sexual differentiation in Drosophila. Female and male isoforms (DSX F and DSX M) are formed due to sex-specific RNA splicing. DNA recognition, mediated by a shared N-terminal zinc module (the DM domain), is enhanced by a C-terminal dimerization element. Sex-specific extension of this element in DSX F and DSX M leads to assembly of distinct transcriptional preinitiation complexes. Here, we describe the structure of the extended C-terminal dimerization domain of DSX F as determined by multidimensional NMR spectroscopy. The core dimerization element is well ordered, giving rise to a dense network of interresidue nuclear Overhauser enhancements. The structure contains dimer-related UBA folds similar to those defined by x-ray crystallographic studies of a truncated domain. Whereas the proximal portion of the female tail extends helix 3 of the UBA fold, the distal tail is disordered. Ala substitutions in the proximal tail disrupt the sex-specific binding of IX (Intersex), an obligatory partner protein and putative transcriptional coactivator; IX-DSX F interaction is, by contrast, not disrupted by truncation of the distal tail. Mutagenesis of the UBA-like dimer of DSX F highlights the importance of steric and electrostatic complementarity across the interface. Two temperature-sensitive mutations at this interface have been characterized in yeast model systems. One weakens a network of solvated salt bridges, whereas the other perturbs the underlying nonpolar interface. These mutations confer graded gene-regulatory activity in yeast within a physiological temperature range and so may provide novel probes for genetic analysis of a sex-specific transcriptional program in Drosophila development.
Yang
Yanwu
Y
Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106, USA.
Zhang
Wei
W
Bayrer
James R
JR
Weiss
Michael A
MA
D000818:Animals; D018360:Crystallography, X-Ray / Q000379:methods; D004268:DNA-Binding Proteins / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics*; D004331:Drosophila melanogaster; D005260:Female; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D008957:Models, Genetic; D008968:Molecular Conformation; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017434:Protein Structure, Tertiary; D012727:Sex Characteristics; D012737:Sex Factors; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; C070369:DSX protein, Drosophila; D029721:Drosophila Proteins