The genome of eukaryotic cells is packaged into chromatin. Recent technological advances have allowed the profiling of chromatin on a genome-wide scale in a diversity of living species. We are now faced with an excellent opportunity to study chromatin function and evolution in a way that was not possible before. Here, I propose to study the mechanisms of nucleosome retention in human sperm and the role and evolution of chromatin architecture at regulatory elements of animal genomes. In human sperm, most of the genome is packaged by protamines and not by histones. The 4% of the human genome that remains packaged in nucleosomes tends to contain gene promoters and developmental regulatory genes. However, there is no mechanism to explain why nucleosomes are retained at these locations. To this end, I will test three possible mechanisms of nucleosome retention in human sperm. This information will help us understand the mechanisms that may affect epigenetic inheritance. As a second aspect of this project, I propose to study the interplay between chromatin structure and regulatory evolution in animal genomes. Nucleosome positions have been mapped throughout five entire animal genomes; human, mouse, Japanese killifish (Oryzias latipes), Caenorhabditis elegans and Drosophila melanogaster. I propose to use this data and the intrinsic affinity for nucleosomes predicted from in vitro nucleosome reconstitution experiments to study the role of nucleosome positioning in the evolution of promoters and distal enhancers in metazoans. The grant will be used for the transition from a post-doctoral to an independent research position and will contribute to my integration in a major scientific hub in Europe.