Split cGAL, an intersectional strategy using a split intein for refined spatiotemporal transgene control in Caenorhabditis elegans.
Proc Natl Acad Sci U S A
; 115(15): 3900-3905, 2018 04 10.
Article
em En
| MEDLINE
| ID: mdl-29581308
ABSTRACT
Bipartite expression systems, such as the GAL4-UAS system, allow fine manipulation of gene expression and are powerful tools for interrogating gene function. Recently, we established cGAL, a GAL4-based bipartite expression system for transgene control in Caenorhabditis elegans, where a single promoter dictates the expression pattern of a cGAL driver, which then binds target upstream activation sequences to drive expression of a downstream effector gene. Here, we report a split strategy for cGAL using the split intein gp41-1 for intersectional control of transgene expression. Split inteins are protein domains that associate, self-excise, and covalently ligate their flanking peptides together. We split the DNA binding domain and transcriptional activation domain of cGAL and fused them to the N terminal of gp41-1-N-intein and the C terminal of gp41-1-C-intein, respectively. In cells where both halves of cGAL are expressed, a functional cGAL driver is reconstituted via intein-mediated protein splicing. This reconstitution allows expression of the driver to be dictated by two promoters for refined spatial control or spatiotemporal control of transgene expression. We apply the split cGAL system to genetically access the single pair of MC neurons (previously inaccessible with a single promoter), and reveal an important role of protein kinase A in rhythmic pharyngeal pumping in C. elegans Thus, the split cGAL system gives researchers a greater degree of spatiotemporal control over transgene expression, and will be a valuable genetic tool in C. elegans for dissecting gene function with finer cell-specific resolution.
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Assunto principal:
Proteínas Fúngicas
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Regulação da Expressão Gênica
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Caenorhabditis elegans
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Transgenes
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Inteínas
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Proteínas de Ligação a DNA
Limite:
Animals
Idioma:
En
Revista:
Proc Natl Acad Sci U S A
Ano de publicação:
2018
Tipo de documento:
Article