Drosophila Brakeless Interacts with Atrophin and Is Required for Tailless-Mediated Transcriptional Repression in Early Embryos

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Abstract
Complex gene expression patterns in animal development are generated by the interplay of transcriptional activators and repressors at cis-regulatory DNA modules (CRMs). How repressors work is not well understood, but often involves interactions with co-repressors. We isolated mutations in the brakeless gene in a screen for maternal factors affecting segmentation of the Drosophila embryo. Brakeless, also known as Scribbler, or Master of thickveins, is a nuclear protein of unknown function. In brakeless embryos, we noted an expanded expression pattern of the Krüppel (Kr) and knirps (kni) genes. We found that Tailless-mediated repression of kni expression is impaired in brakeless mutants. Tailless and Brakeless bind each other in vitro and interact genetically. Brakeless is recruited to the Kr and kni CRMs, and represses transcription when tethered to DNA. This suggests that Brakeless is a novel co-repressor. Orphan nuclear receptors of the Tailless type also interact with Atrophin co-repressors. We show that both Drosophila and human Brakeless and Atrophin interact in vitro, and propose that they act together as a co-repressor complex in many developmental contexts. We discuss the possibility that human Brakeless homologs may influence the toxicity of polyglutamine-expanded Atrophin-1, which causes the human neurodegenerative disease dentatorubral-pallidoluysian atrophy (DRPLA). Nuclear receptors play important roles in embryonic development and cellular differentiation by regulating gene expression at the level of transcription. The functions of transcriptional repressors, including nuclear receptors, are often mediated by other proteins, so-called co-repressors. We performed a genetic screen in the fruit fly Drosophila melanogaster to search for novel co-repressor proteins. We isolated mutations in the brakeless gene that alter normal transcriptional repression in early fly embryos. Brakeless was already known to regulate axon guidance in the eye, larval behavior, and gene expression in wing imaginal discs. However, the molecular function of this protein was unknown. Here we show that Brakeless is a co-repressor required for function of the Tailless nuclear receptor. Tailless was previously shown to interact with another co-repressor, Atrophin. Here, we demonstrate that Brakeless and Atrophin can bind to one another and that this interaction is conserved between a human Brakeless homolog, ZNF608, and human Atrophin-1. A polyglutamine expansion in Atrophin-1 is the cause of the neurodegenerative disease dentatorubral-pallidoluysian atrophy (DRPLA). It is possible that the interaction with ZNF608 could contribute to the pathogenesis of polyglutamine-expanded Atrophin-1.