The role of histone chaperones and specific transcription factors during multicellular development of filamentous ascomycetes

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The aim of this project is the molecular analysis of fruiting body development in filamentous fungi. Fruiting body development is an ontogenetic program that is an essential part of the life cycle of many filamentous fungi; however, its molecular basis is far from understood. In the proposed project we will analyze the role of factors that regulate gene expression at the level of transcription in the model organism Sordaria macrospora. The analyses will be performed with omics-techniques to obtain a genome-wide overview of nuclear processes that occur during fruiting body development. In previous analyses, several transcription factors and histone chaperones involved in fruiting body development were identified. Furthermore, we have established a combination of laser microdissection and RNA-seq to study the organ-specific gene expression in young fruiting bodies of the wild type and the developmental mutant pro1 that carries a mutation in a transcription factor gene. These analyses will be extended to include other transcription factor and histone chaperone mutants to unravel the regulatory network that coordinates chromatin structure and transcription during fruiting body development. In addition, we will establish ChIP-seq to identify genomic regions that are bound by transcription factors. A comparison of RNA-seq and ChIP-seq data can be used to distinguish between direct and indirect targets of transcription factors. We will also include histone chaperones in the ChIP-seq analyses; this requires crosslinking of histone chaperones, histones, and DNA prior to the chromatin immunoprecipitation, because the histone chaperones are not directly binding to DNA. These techniques will be used to analyze the influence of histone chaperones and transcription factors in parallel, which allows the identification of interdependencies. A second aspect of the project is the identification of proteins that interact with the histone chaperones and transcription factors under investigation. We have already identified interaction partners of the histone chaperone ASF1 using a highly sensitive mass spectrometry (MudPIT), and these analyses will be performed for the other developmental factors. The combination of several high-throughput techniques for the analysis of gene expression will yield an integrated picture with a high temporal and spatial resolution of the molecular processes that are orchestrate fruiting body development.