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Methods. 2014 Jun 15;68(1):194-8. doi: 10.1016/j.ymeth.2014.03.015. Epub 2014 Mar 25.

Modulating and measuring Wingless signalling.

Author information

1
MRC National Institute for Medical Research, The Ridgeway, Mill Hill, London NW71AA, United Kingdom. Electronic address: jvincen@nimr.mrc.ac.uk.

Abstract

The main Wnt ligand of Drosophila activates a conserved canonical signalling pathway to regulate a plethora of cellular activities during development, regeneration and nervous system function. Here I first describe experimental means of measuring and modulating Wingless signalling in Drosophila cell culture. Various reporters have been devised by placing TCF-binding sites or DNA fragments from known target genes upstream of luciferase-coding sequences. Signalling can be activated in cells by addition of Wingless conditioned medium, treatment with a chemical inhibitor of Shaggy/GSK3 or transfection with a plasmid encoding activated Armadillo (Drosophila β-catenin). Measuring Wingless signalling in intact tissue is somewhat more challenging than in cell culture. Synthetic transgenic reporters have been devised but further improvements are needed to achieve sensitive responsiveness to Wingless at all times and places. As an alternative, gene traps in frizzled3 and notum/wingful, two context-independent endogenous targets, can be used as reporters. It is hoped that further modification of these loci could lead to more versatile and sensitive means of detecting signalling. Many genetic tools are available to trigger ectopic signalling or prevent endogenous signalling. These mostly rely on RNAi-producing transgenes or the generation of mutant patches by mitotic recombination. New developments in genome engineering are opening further means of manipulating the components of Wingless signalling with exquisite temporal and spatial precision.

KEYWORDS:

Drosophila; Reporter gene; Signalling; Wingless

PMID:
24675402
DOI:
10.1016/j.ymeth.2014.03.015
[Indexed for MEDLINE]

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