Genome editing enables reverse genetics of multicellular development in the choanoflagellate Salpingoeca rosetta

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Abstract

In a previous study, we established a forward genetic screen to identify genes required for multicellular development in the choanoflagellate, Salpingoeca rosetta (Levin et al., 2014). Yet, the paucity of reverse genetic tools for choanoflagellates has hampered direct tests of gene function and impeded the establishment of choanoflagellates as a model for reconstructing the origin of their closest living relatives, the animals. Here we establish CRISPR/Cas9-mediated genome editing in S. rosetta by engineering a selectable marker to enrich for edited cells. We then use genome editing to disrupt the coding sequence of a S. rosetta C-type lectin gene, rosetteless, and thereby demonstrate its necessity for multicellular rosette development. This work advances S. rosetta as a model system in which to investigate how genes identified from genetic screens and genomic surveys function in choanoflagellates and evolved as critical regulators of animal biology.

Data availability

All data generated are included in the manuscript. Additionally, we have posted a protocol at protocols.io: https://dx.doi.org/10.17504/protocols.io.89fhz3n

Article and author information

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David S Booth

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

For correspondence
David.Booth@ucsf.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-4724-4702
Nicole King

Department of Molecular and Cell Biology, Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-6409-1111

Funding

Gordon and Betty Moore Foundation

David S Booth
Nicole King

Howard Hughes Medical Institute

Nicole King

Jane Coffin Childs Memorial Fund for Medical Research

David S Booth

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.