Cre-assisted fine-mapping of neural circuits using orthogonal split inteins

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Abstract

Existing genetic methods of neuronal targeting do not routinely achieve the resolution required for mapping brain circuits. New approaches are thus necessary. Here, we introduce a method for refined neuronal targeting that can be applied iteratively. Restriction achieved at the first step can be further refined in a second step, if necessary. The method relies on first isolating neurons within a targeted group (i.e. Gal4 pattern) according to their developmental lineages, and then intersectionally limiting the number of lineages by selecting only those in which two distinct neuroblast enhancers are active. The neuroblast enhancers drive expression of split Cre recombinase fragments. These are fused to non-interacting pairs of split inteins, which ensure reconstitution of active Cre when all fragments are expressed in the same neuroblast. Active Cre renders all neuroblast-derived cells in a lineage permissive for Gal4 activity. We demonstrate how this system can facilitate neural circuit-mapping in Drosophila.

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Data generated during this study are included in the manuscript and supporting files. Source data files have been provided for Figure 2I

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Haojiang Luan

Laboratory of Molecular Biology, National Institute of Mental Health, Bethesda, United States

Competing interests
The authors declare that no competing interests exist.
Alexander Kuzin

Neural Cell-Fate Determinants Section, National Institute of Neurological Disease and Stroke, Bethesda, United States

Competing interests
The authors declare that no competing interests exist.
Ward F Odenwald

Neural Cell-Fate Determinants Section, National Institute of Neurological Disease and Stroke, Bethesda, United States

Competing interests
The authors declare that no competing interests exist.
Benjamin H White

Laboratory of Molecular Biology, National Institute of Mental Health, Bethesda, United States

For correspondence
benjaminwhite@mail.nih.gov

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0003-0612-8075

Funding

National Institute of Mental Health (ZIA-MH002800)

Benjamin H White

National Institute of Neurological Disorders and Stroke (ZIA-NS002820-26)

Ward F Odenwald

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

Copyright

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.