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Deciphering the mechanisms that regulate gene expression is central to understanding life at the molecular level. The advent of Big Data in biology, and the implementation of techniques for single molecule analyses at many different levels have opened new horizons for the study of transcription regulation. Still, the unicellular yeast remains one of the best systems to integrate analyses from multidisciplinary research areas and the findings are often transposable to more complex higher eukaryotes. This EMBO Workshop is unique as it brings together scientists working on very different aspects of gene expression with a variety of approaches including genomics, structural biology, systems analyses and computational modeling. Many topics will be addressed, including chromatin dynamics, mechanisms of transcription, the regulatory role of non-coding transcription and RNA metabolism either globally or in single cells.