The canonical model of gene regulation is based on enhancer-promoter interactions to determine spatiotemporal properties of gene expression. So far, most studies on gene regulation in complex multicellular organisms have focused on particular tissues or cell types, having limited power in investigating possible higher hierarchical level gene regulatory principles. Therefore, we set out to investigate the rules of gene regulation and enhancer grammar in the context of a whole organism using sequence-to-function models. For this, we generated a whole adult fly scATAC-seq atlas containing around 700,000 cells and 150,000 peaks that are accessible in at least one cell type. We used topic modelling to cluster the cells in the atlas and transferred cell type annotations from the scRNA-seq atlas at two hierarchical levels (class and specific). This resulted in a pseudo-multiome atlas with matched scATAC-seq and scRNA-seq profiles across more than 100 cell types. We stratified scATAC-seq peaks according to their accessibility across cell types into ubiquitous, cell-type-specific, and putative cell-class-specific types. Next, we trained a suite of local and global sequence-to-function models to understand the grammars of the three classes of regions we identified and to integrate locus level information to predict gene regulation. Using these models, we identified and validated cell-type-specific enhancers for different cell types, investigated the promoters of genes and the relation of promoter-related motifs to gene expression patterns, and investigated the grammar and gene expression effects of a novel putative class of regions that we call 'class-level enhancers'.

Co-authors: Anton De Brabandere, Swann Floc'Hlay, Berfin Dag, Valerie Christiaens, Katina Spanier, Stein Aerts