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Genomes are arranged in a confined space in the cell, the nucleoid or nucleus. This arrangement is hierarchical and dynamic, and follows DNA/chromatin-based transactions or environmental conditions. Describing the interplay between local genome structure and gene activity is a long-standing quest in biology. Here, we focus on systematic studies correlating bacterial genome folding and function. Parallels on organizational similarities with eukaryotes are drawn. The biological relevance of hierarchical units in bacterial genome folding and the causal relationship between genome folding and its activity is unclear. We discuss recent quantitative approaches to tackle these questions. Moreover, we sketch a perspective of experiments necessary to iteratively and systematically build, test and improve structure–function models of bacterial chromatin.