Functional enhancer elements drive subclass-selective expression from mouse to primate neocortex

Viral genetictools that target specificbraincell types could transform basicneuroscienceand targeted gene therapy. Here, we use comparative open chromatin analysis to identify thousands of human-neocortical-subclass-specific putative enhancers from across the genome tocontrol gene expressionin adeno-associated virus (AAV) vectors. The cellular specificity of reporter expression from enhancer-AAVs is established by molecular profiling after systemic AAV delivery in mouse. Over 30% of enhancer-AAVs produce specific expression in the targeted subclass, including both excitatory and inhibitory subclasses. We present a collection ofParvalbumin(PVALB) enhancer-AAVs that show highly enriched expression not only in cortical PVALB cells but also in some subcortical PVALB populations. Five vectors maintain PVALB-enriched expression in primateneocortex. These results demonstrate how genome-wide open chromatin data mining and cross-species AAV validation can be used to create the next generation of non-species-restricted viral genetic tools. Single-cell RNA-seq on flow-sorted enhancer-AAV-labeled transgene-expressing cells from 33 mouse brains. From each mouse brain up to 48 individual cells were profiled by SMARTer single-cell RNA-seq. These data are part of the study located at https://www.ncbi.nlm.nih.gov/projects/gap/cgi-bin/study.cgi?study_id=phs002292.v1.p1.