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Use of a molecular genetic platform technology to produce human Wnt proteins reveals distinct local and distal signaling abilities.

ABSTRACT: Functional and mechanistic studies of Wnt signaling have been severely hindered by the inaccessibility of bioactive proteins. To overcome this long-standing barrier, we engineered and characterized a panel of Chinese hamster ovary (CHO) cell lines with inducible transgenes encoding tagged and un-tagged human WNT1, WNT3A, WNT5A, WNT7A, WNT11, WNT16 or the soluble Wnt antagonist Fzd8CRD, all integrated into an identical genomic locus. Using a quantitative real-time bioluminescence assay, we show that cells expressing WNT1, 3A and 7A stimulate Wnt/beta-catenin reporter activity, while the other WNT expressing cell lines interfere with this activation. Additionally, in contrast to WNT3A, WNT1 only exhibits activity when cell-associated, and thus only signals to neighboring cells. The reporter assay also revealed a rapid decline of Wnt activity at 37°C, indicating that Wnt activity is highly labile. These engineered cell lines will reduce the cost of making and purifying Wnt proteins and serve as a continuous, reliable and regulatable source of Wnts to research laboratories around the world.

SUBMITTER: Green JL

PROVIDER: S-EPMC3596392 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Use of a molecular genetic platform technology to produce human Wnt proteins reveals distinct local and distal signaling abilities.

Green Jennifer L JL Bauer Matthieu M Yum Kyu Won KW Li Yao-Cheng YC Cox Miranda L ML Willert Karl K Wahl Geoffrey M GM

PloS one 20130313 3

Functional and mechanistic studies of Wnt signaling have been severely hindered by the inaccessibility of bioactive proteins. To overcome this long-standing barrier, we engineered and characterized a panel of Chinese hamster ovary (CHO) cell lines with inducible transgenes encoding tagged and un-tagged human WNT1, WNT3A, WNT5A, WNT7A, WNT11, WNT16 or the soluble Wnt antagonist Fzd8CRD, all integrated into an identical genomic locus. Using a quantitative real-time bioluminescence assay, we show t ...[more]

PMID: 23516471

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Use of a molecular genetic platform technology to produce human Wnt proteins reveals distinct local and distal signaling abilities.

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Use of a molecular genetic platform technology to produce human Wnt proteins reveals distinct local and distal signaling abilities.

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