The Doublesex transcription factor: Structural and functional studies of a sex-determining factor

 
 
 
 

The Doublesex transcription factor: Structural and functional studies of a sex-determining factor

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Title: The Doublesex transcription factor: Structural and functional studies of a sex-determining factor
Author: Bayrer, James Robert
Description: Doublesex (DSX) is a transcription factor responsible for the regulation of sexual differentiation in Drosophila. Alternate splicing gives rise to male- and female-specific isoforms. A potent modulator of the yolk protein gene (yp), the male isoform (DSXM) represses transcription of yp whereas the female isoform (DSXF) is an activator. DSX contains two recognized domains, an N-terminal DNA-binding domain (shared between isoforms) and a C-terminal domain (CTD) that is responsible for oligomerization and presumably transcriptional regulation. The CTDs contain sex-specific C-terminal sequences with opposite gene-regulatory properties. We have solved the crystal structure of the core CTD dimerization domain to a resolution of 1.6 Å using single-wavelength anomalous dispersion (SAD) phasing. The crystal structure reveals a novel dimeric arrangement of ubiquitin-associated (UBA) folds. To our knowledge this is its first report in a transcription factor, and the first structure of a dimeric UBA domain. Dimerization is mediated by a non-canonical hydrophobic interface extrinsic to the putative Ub-binding surface. The unexpected observation of a UBA fold in DSX extends the repertoire of -helical dimerization elements in transcription factors. Intersexual development of XX Drosophila (karyotypic females) is associated with mutation G398D, encoded in female-specific exon 4. G398 lies within CTD. We demonstrate that the intersexual mutation blocks dimerization leading to non-native aggregation. Analysis of diverse substitutions at 398 indicates that only alanine (found in the male isoform) is tolerated. The structure of the dimer suggests that side chains larger than alanine would clash at the interface; dimerization of the G398D variant would be further destabilized by an uncompensated buried charge. The instability of the variant monomer suggests that folding and dimerization are coupled. Such coupling ― although a general feature of leucine zippers and other helical dimerization elements ― is novel among UBA domains. We extend these studies by hydrogen exchange analysis, demonstrating long-lived resonances in the dimer interface. We further present evidence for ubiquitin-binding by CTDF, suggesting a previously unappreciated role for ubiquitin in sex determination. Our results define a new role for the UBA fold in transcription and rationalize the impaired sexual differentiation of a model organism.
Permanent Link: http://rave.ohiolink.edu/etdc/view?acc_num=case1124911161
http://hdl.handle.net/2374.OX/17352
Date: 2006

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