SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation
Background: Conserved Wnt ligands are critical for signalling during development; however, various factors modulate their activity. Among these factors are the Secreted Frizzled-Related Proteins (SFRP). We previously isolated the SFRP-4 gene from an involuting rat mammary gland and later showed that...
| Main Authors: | , , , , , |
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| Format: | Journal Article |
| Published: |
2008
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| Online Access: | http://hdl.handle.net/20.500.11937/25446 |
| _version_ | 1848751711334694912 |
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| author | Constantinou, T. Baumann, F. Lacher, M. Saurer, S. Friis, R. Dharmarajan, Arunasalam |
| author_facet | Constantinou, T. Baumann, F. Lacher, M. Saurer, S. Friis, R. Dharmarajan, Arunasalam |
| author_sort | Constantinou, T. |
| building | Curtin Institutional Repository |
| collection | Online Access |
| description | Background: Conserved Wnt ligands are critical for signalling during development; however, various factors modulate their activity. Among these factors are the Secreted Frizzled-Related Proteins (SFRP). We previously isolated the SFRP-4 gene from an involuting rat mammary gland and later showed that transgenic mice inappropriately expressing SFRP-4 during lactation exhibited a high level of apoptosis with reduced survival of progeny. Results: In order to address the questions related to the mechanism of Wnt signalling and its inhibition by SFRP-4 which we report here, we employed partially-purified Wnt-3a in a co-culture model system. Ectopic expression of SFRP-4 was accomplished by infection with a pBabepuro construct. The co-cultures comprised Line 31E mouse mammary secretory epithelial cells and Line 30F, undifferentiated, fibroblast-like mouse mammary cells. In vitro differentiation of such co-cultures can be demonstrated by induction of the ß-casein gene in response to lactogenic hormones. We show here that treatment of cells with partially-purified Wnt-3a initiates Dvl-3, Akt/PKB and GSK-3ß hyperphosphorylation and ß-catenin activation. Furthermore, while up-regulating the cyclin D1 and connexin-43 genes and elevating transepithelial resistance of Line 31E cell monolayers, Wnt-3a treatment abrogates differentiation of co-cultures in response to the lactogenic hormones prolactin, insulin and glucocorticoid. Cells which express SFRP-4, however, are largely unaffected by Wnt-3a stimulation. Since a physical association between Wnt-3a and SFRP-4 could be demonstrated with immunoprecipitation/Western blotting experiments, this interaction, presumably owing to the Frizzled homology region typical of all SFRPs, explains the refractory response to Wnt-3a which was observed. Conclusion: This study demonstrates that Wnt-3a treatment activates the Wnt signalling pathway and interferes with in vitro differentiation of mammary co-cultures to ß-casein production in response to lactogenic hormones. Similarly, in another measure of differentiation, following Wnt-3a treatment mammary epithelial cells could be shown to up-regulate the cyclin D1 and connexin-43 genes while phenotypically they show increased transepithelial resistance across the cell monolayer. All these behavioural changes can be blocked in mammary epithelial cells expressing SFRP-4. Thus, our data illustrate in an in vitro model a mechanism by which SFRP-4 can modulate a differentiation response to Wnt-3a. © 2008 Constantinou et al; licensee BioMed Central Ltd. |
| first_indexed | 2025-11-14T07:57:04Z |
| format | Journal Article |
| id | curtin-20.500.11937-25446 |
| institution | Curtin University Malaysia |
| institution_category | Local University |
| last_indexed | 2025-11-14T07:57:04Z |
| publishDate | 2008 |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | curtin-20.500.11937-254462017-09-13T15:17:39Z SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation Constantinou, T. Baumann, F. Lacher, M. Saurer, S. Friis, R. Dharmarajan, Arunasalam Background: Conserved Wnt ligands are critical for signalling during development; however, various factors modulate their activity. Among these factors are the Secreted Frizzled-Related Proteins (SFRP). We previously isolated the SFRP-4 gene from an involuting rat mammary gland and later showed that transgenic mice inappropriately expressing SFRP-4 during lactation exhibited a high level of apoptosis with reduced survival of progeny. Results: In order to address the questions related to the mechanism of Wnt signalling and its inhibition by SFRP-4 which we report here, we employed partially-purified Wnt-3a in a co-culture model system. Ectopic expression of SFRP-4 was accomplished by infection with a pBabepuro construct. The co-cultures comprised Line 31E mouse mammary secretory epithelial cells and Line 30F, undifferentiated, fibroblast-like mouse mammary cells. In vitro differentiation of such co-cultures can be demonstrated by induction of the ß-casein gene in response to lactogenic hormones. We show here that treatment of cells with partially-purified Wnt-3a initiates Dvl-3, Akt/PKB and GSK-3ß hyperphosphorylation and ß-catenin activation. Furthermore, while up-regulating the cyclin D1 and connexin-43 genes and elevating transepithelial resistance of Line 31E cell monolayers, Wnt-3a treatment abrogates differentiation of co-cultures in response to the lactogenic hormones prolactin, insulin and glucocorticoid. Cells which express SFRP-4, however, are largely unaffected by Wnt-3a stimulation. Since a physical association between Wnt-3a and SFRP-4 could be demonstrated with immunoprecipitation/Western blotting experiments, this interaction, presumably owing to the Frizzled homology region typical of all SFRPs, explains the refractory response to Wnt-3a which was observed. Conclusion: This study demonstrates that Wnt-3a treatment activates the Wnt signalling pathway and interferes with in vitro differentiation of mammary co-cultures to ß-casein production in response to lactogenic hormones. Similarly, in another measure of differentiation, following Wnt-3a treatment mammary epithelial cells could be shown to up-regulate the cyclin D1 and connexin-43 genes while phenotypically they show increased transepithelial resistance across the cell monolayer. All these behavioural changes can be blocked in mammary epithelial cells expressing SFRP-4. Thus, our data illustrate in an in vitro model a mechanism by which SFRP-4 can modulate a differentiation response to Wnt-3a. © 2008 Constantinou et al; licensee BioMed Central Ltd. 2008 Journal Article http://hdl.handle.net/20.500.11937/25446 10.1186/1750-2187-3-10 unknown |
| spellingShingle | Constantinou, T. Baumann, F. Lacher, M. Saurer, S. Friis, R. Dharmarajan, Arunasalam SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation |
| title | SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation |
| title_full | SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation |
| title_fullStr | SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation |
| title_full_unstemmed | SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation |
| title_short | SFRP-4 abrogates Wnt-3a-induced ß-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation |
| title_sort | sfrp-4 abrogates wnt-3a-induced ß-catenin and akt/pkb signalling and reverses a wnt-3a-imposed inhibition of in vitro mammary differentiation |
| url | http://hdl.handle.net/20.500.11937/25446 |