ZIP of imaging-based data files.

Immune responses in tissues display complex spatial patterns of gene expression that are linked to disease outcomes. However, the processes that generate these patterns—including the relative roles of noisy gene expression dynamics, microbial transport, and tissue anatomy—are poorly understood. As a tractable model of spatial immune responses, we investigated heterogeneous expression of antimicrobial peptides in the larval fly fat body, an organ functionally analogous to the liver. To quantify single-cell antimicrobial peptide expression dynamics in the fat body, we developed a protocol for light sheet fluorescence microscopy of whole, live larvae. Using this approach, we discovered that individual fat body cells express antimicrobial peptides at approximately constant rates following infection, but that the average rate varies along the anterior-posterior axis of the fat body, with rapid expression in the anterior and posterior lobes. Overexpression of immune signaling components and analysis of spatial transcriptomes revealed that these tissue microenvironments are predefined independently of infection, with the rate-limiting step of antimicrobial peptide induction downstream of peptidoglycan sensing. The locations of these microevironments correlate with heartbeat-dependent fluid flow in a manner resembling the strategic positioning of immune cells in the liver, gut, and lymph nodes of mammals. We speculate that this spatial compartmentalization helps the fat body efficiently perform its diverse metabolic, enzymatic, and immunological functions.