Spatial motifs reveal patterns in cellular architecture of complex tissues

Spatial organization of cells is crucial to both proper physiological function of tissues and pathological conditions like cancer. Recent advances in spatial transcriptomics have enabled joint profiling of gene expression and spatial context of the cells. The outcome is an information rich map of the tissue where individual cells, or small regions, can be labeled based on their gene expression state. While spatial transcriptomics excels in its capacity to profile numerous genes within the same sample, most existing methods for analysis of spatial data only examine distribution of one or two labels at a time. These approaches overlook the potential for identifying higher-order associations between cell types - associations that can play a pivotal role in understanding development and function of complex tissues. In this context, we introduce a novel method for detecting motifs in spatial neighborhood graphs. Each motif represents a spatial arrangement of cell types that occurs in the tissue more frequently than expected by chance. To identify spatial motifs, we developed an algorithm for uniform sampling of paths from neighborhood graphs and combined it with a motif finding algorithm on graphs inspired by previous methods for finding motifs in DNA sequences. Using synthetic data with known ground truth, we show that our method can identify spatial motifs with high accuracy and sensitivity. Applied to spatial maps of mouse retinal bipolar cells and hypothalamic preoptic region, our method reveals previously unrecognized patterns in cell type arrangements. In some cases, cells within these spatial patterns differ in their gene expression from other cells of the same type, providing insights into the functional significance of the spatial motifs. These results suggest that our method can illuminate the substantial complexity of neural tissues, provide novel insight even in well studied models, and generate experimentally testable hypotheses.