Functional characterisation of LINT and other dCoREST complexes

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Mutation of the l(3)mbt gene of Drosophila cause the formation of malignant brain tumours. In order to better understand the molecular functions of L(3)mbt protein, we have purified L(3)mbt-associated polypeptides and identified the L(3)mbt-containing LINT complex. The LINT complex is a molecular machine that consists of three subunits (L(3)mbt, Lint-1 and CoREST). It binds to chromatin and represses the transcription of brain tumour-associated genes. We have determined the genomic binding sites of the complex, have defined its target genes and established assays to investigate its molecular repression mechanisms. During purification of LINT we have discovered additional complexes which also contain a CoREST subunit.During the second funding period we want to address important questions about the LINT complex and we want to functionally characterise the newly discovered CoREST protein complex family. We will reconstitute LINT from recombinant subunits, define domains that are important for complex integrity and determine the interactions between LINT and modified polynucleosomes in vitro. In addition, we will define the parameters that allow LINT to specifically bind to its target genes and we will elucidate its mechanism of gene repression. We will analyse the role of LINT in cell fate specification using the Drosophila wing as a model. In addition, we will characterise the novel CoREST protein complex familiy of Drosophila, by systematically purifying and characterising all CoREST-containing protein complexes.Given that both L(3)mbt and CoREST are conserved in humans, we hope that our studies will shed light on disease-relevant molecular mechanisms.