Perturb-FISH simultaneous CRISPR screening and spatial transcriptomics in astrocytes

[ DOI: https://doi.org/10.35077/ace-gem-get, BIL ID: ace-gem-get ]

Dataset Citation:

Binan, Loϊc, Binan, Loϊc. (2024). Perturb-FISH simultaneous CRISPR screening and spatial transcriptomics in astrocytes [Dataset/Microscopy]. Brain Image Library. https://doi.org/10.35077/ace-gem-get

 

Abstract:

Pooled optical screens have enabled the study of cellular interactions, morphology, or dynamics at massive scale, but have not yet leveraged the power of highly-plexed single-cell resolved transcriptomic readouts to inform molecular pathways. Here, we present Perturb-FISH, which bridges these approaches by combining imaging spatial transcriptomics with parallel optical detection of in situ amplified guide RNAs. We show that Perturb-FISH recovers intracellular effects that are consistent with Perturb-seq results in a screen of lipopolysaccharide response in cultured monocytes, and uncover new intercellular and density-dependent regulation of the innate immune response. We further pair Perturb-FISH with a functional readout in a screen of autism spectrum disorder risk genes, showing common calcium activity phenotypes in induced pluripotent stem cell derived astrocytes and their associated genetic interactions and dysregulated molecular pathways. Perturb-FISH is thus a generally applicable method for studying the genetic and molecular associations of spatial and functional biology at single-cell resolution.

 

Methods:

A full protocol describing the technology introduced in this work is available at protocols.io: https://dx.doi.org/10.17504/protocols.io.6qpvr356bvmk/v1

 

Contributors and Roles:

 

Funding:

 

Technical Information:

spatial transcriptomics (Modality) / MERFISH (Technique)

Computational analysis was performed in two parts: the decoding of the MERFISH signal was performed using the google cloud compute platform, using MERlin: https://github.com/lbinan/gcp_merlin

 

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