Scalable in vivo cardiac functional genomics with compressed AAV-Perturb-seq reveals a common mitochondrial response...

Efficient identification of new targets to treat human disease requires a scalable way to link genotype to phenotype directly in the target organ. Pooled CRISPR screening with single-cell RNA sequencing as a readout (Perturb-seq) has emerged as a method for functional genomics but is typically applied in vitro and is limited in scale. Here, we combine in vivo Perturb-seq via adeno-associated virus (AAV)-mediated delivery with a statistical framework allowing for signal deconvolution after multiple random perturbations per cell (compressed Perturb-seq), to develop in vivo compressed AAV-Perturb-seq, a scalable way to perform high-throughput, cell-autonomous functional genomics in a desired target organ. We apply this approach to study the effect of 585 gene knockouts on the cardiomyocyte transcriptome. We identify that alterations in the mitochondrial transcriptome are a common response to genetic perturbation, a finding independently validated across species and perturbation modalities. We identify very few synergistic perturbations, despite observing frequent combinatorial effects. Broadly, our work establishes a platform to facilitate in vivo functional genomics in a target organ with direct applicability to identifying therapeutic targets for the treatment of human disease.