PT - JOURNAL ARTICLE AU - Hans-Hermann Wessels AU - Alejandro Méndez-Mancilla AU - Efthymia Papalexi AU - William M Mauck AU - Lu Lu AU - John A. Morris AU - Eleni Mimitou AU - Peter Smibert AU - Neville E. Sanjana AU - Rahul Satija TI - Efficient combinatorial targeting of RNA transcripts in single cells with Cas13 RNA Perturb-seq AID - 10.1101/2022.02.02.478894 DP - 2022 Jan 01 TA - bioRxiv PG - 2022.02.02.478894 4099 - http://biorxiv.org/content/early/2022/02/02/2022.02.02.478894.short 4100 - http://biorxiv.org/content/early/2022/02/02/2022.02.02.478894.full AB - Pooled CRISPR screens coupled with single-cell RNA-sequencing have enabled systematic interrogation of gene function and regulatory networks. Here, we introduce Cas13 RNA Perturb-seq (CaRPool-seq) which leverages the RNA-targeting CRISPR/Cas13d system and enables efficient combinatorial perturbations alongside multimodal single-cell profiling. CaRPool-seq encodes multiple perturbations on a cleavable array which is associated with a detectable barcode sequence, allowing for the simultaneous targeting of multiple genes. We compared CaRPool-seq to existing Cas9-based methods, highlighting its unique strength to efficiently profile combinatorially perturbed cells. Finally, we apply CaRPool-seq to perform multiplexed combinatorial perturbations of myeloid differentiation regulators in an acute myeloid leukemia (AML) model system and identify extensive interactions between different chromatin regulators that can enhance or suppress AML differentiation phenotypes.Competing Interest StatementIn the past three years, RS has worked as a consultant for Bristol-Myers Squibb, Regeneron, and Kallyope, and served as a SAB member for ImmunAI, Apollo Life Sciences GmbH, Nanostring, and the NYC Pandemic Response Lab. NES is an advisor to Vertex. PS is a co-inventor on a patent related to protein detection by sequencing as described in this work. The New York Genome Center and New York University have applied for patents relating to the work in this article.