L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness
Abstract Background Metastasis is the predominant cause for cancer morbidity and mortality accounting for approximatively 90% of cancer deaths. The actin-bundling protein L-plastin has been proposed as a metastatic marker and phosphorylation on its residue Ser5 is known to increase its actin-bundlin...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2021-02-01
|
| Series: | Cell Communication and Signaling |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12964-021-00710-5 |
| _version_ | 1818409691649146880 |
|---|---|
| author | Raquel A. C. Machado Dunja Stojevski Sébastien De Landtsheer Philippe Lucarelli Alexandre Baron Thomas Sauter Elisabeth Schaffner-Reckinger |
| author_facet | Raquel A. C. Machado Dunja Stojevski Sébastien De Landtsheer Philippe Lucarelli Alexandre Baron Thomas Sauter Elisabeth Schaffner-Reckinger |
| author_sort | Raquel A. C. Machado |
| collection | DOAJ |
| description | Abstract Background Metastasis is the predominant cause for cancer morbidity and mortality accounting for approximatively 90% of cancer deaths. The actin-bundling protein L-plastin has been proposed as a metastatic marker and phosphorylation on its residue Ser5 is known to increase its actin-bundling activity. We recently showed that activation of the ERK/MAPK signalling pathway leads to L-plastin Ser5 phosphorylation and that the downstream kinases RSK1 and RSK2 are able to directly phosphorylate Ser5. Here we investigate the involvement of the PI3K pathway in L-plastin Ser5 phosphorylation and the functional effect of this phosphorylation event in breast cancer cells. Methods To unravel the signal transduction network upstream of L-plastin Ser5 phosphorylation, we performed computational modelling based on immunoblot analysis data, followed by experimental validation through inhibition/overexpression studies and in vitro kinase assays. To assess the functional impact of L-plastin expression/Ser5 phosphorylation in breast cancer cells, we either silenced L-plastin in cell lines initially expressing endogenous L-plastin or neoexpressed L-plastin wild type and phosphovariants in cell lines devoid of endogenous L-plastin. The established cell lines were used for cell biology experiments and confocal microscopy analysis. Results Our modelling approach revealed that, in addition to the ERK/MAPK pathway and depending on the cellular context, the PI3K pathway contributes to L-plastin Ser5 phosphorylation through its downstream kinase SGK3. The results of the transwell invasion/migration assays showed that shRNA-mediated knockdown of L-plastin in BT-20 or HCC38 cells significantly reduced cell invasion, whereas stable expression of the phosphomimetic L-plastin Ser5Glu variant led to increased migration and invasion of BT-549 and MDA-MB-231 cells. Finally, confocal image analysis combined with zymography experiments and gelatin degradation assays provided evidence that L-plastin Ser5 phosphorylation promotes L-plastin recruitment to invadopodia, MMP-9 activity and concomitant extracellular matrix degradation. Conclusion Altogether, our results demonstrate that L-plastin Ser5 phosphorylation increases breast cancer cell invasiveness. Being a downstream molecule of both ERK/MAPK and PI3K/SGK pathways, L-plastin is proposed here as a potential target for therapeutic approaches that are aimed at blocking dysregulated signalling outcome of both pathways and, thus, at impairing cancer cell invasion and metastasis formation. Video abstract |
| first_indexed | 2024-12-14T10:03:39Z |
| format | Article |
| id | doaj.art-cd6995edc977404c97f35efa1614e4e2 |
| institution | Directory Open Access Journal |
| issn | 1478-811X |
| language | English |
| last_indexed | 2024-12-14T10:03:39Z |
| publishDate | 2021-02-01 |
| publisher | BMC |
| record_format | Article |
| series | Cell Communication and Signaling |
| spelling | doaj.art-cd6995edc977404c97f35efa1614e4e22022-12-21T23:07:11ZengBMCCell Communication and Signaling1478-811X2021-02-0119112210.1186/s12964-021-00710-5L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasivenessRaquel A. C. Machado0Dunja Stojevski1Sébastien De Landtsheer2Philippe Lucarelli3Alexandre Baron4Thomas Sauter5Elisabeth Schaffner-Reckinger6Cancer Cell Biology and Drug Discovery Group, Department of Life Sciences and Medicine, Faculty of Science, Technology and Medicine, University of LuxembourgCancer Cell Biology and Drug Discovery Group, Department of Life Sciences and Medicine, Faculty of Science, Technology and Medicine, University of LuxembourgSystems Biology Group, Department of Life Sciences and Medicine, University of LuxembourgSystems Biology Group, Department of Life Sciences and Medicine, University of LuxembourgCancer Cell Biology and Drug Discovery Group, Department of Life Sciences and Medicine, Faculty of Science, Technology and Medicine, University of LuxembourgSystems Biology Group, Department of Life Sciences and Medicine, University of LuxembourgCancer Cell Biology and Drug Discovery Group, Department of Life Sciences and Medicine, Faculty of Science, Technology and Medicine, University of LuxembourgAbstract Background Metastasis is the predominant cause for cancer morbidity and mortality accounting for approximatively 90% of cancer deaths. The actin-bundling protein L-plastin has been proposed as a metastatic marker and phosphorylation on its residue Ser5 is known to increase its actin-bundling activity. We recently showed that activation of the ERK/MAPK signalling pathway leads to L-plastin Ser5 phosphorylation and that the downstream kinases RSK1 and RSK2 are able to directly phosphorylate Ser5. Here we investigate the involvement of the PI3K pathway in L-plastin Ser5 phosphorylation and the functional effect of this phosphorylation event in breast cancer cells. Methods To unravel the signal transduction network upstream of L-plastin Ser5 phosphorylation, we performed computational modelling based on immunoblot analysis data, followed by experimental validation through inhibition/overexpression studies and in vitro kinase assays. To assess the functional impact of L-plastin expression/Ser5 phosphorylation in breast cancer cells, we either silenced L-plastin in cell lines initially expressing endogenous L-plastin or neoexpressed L-plastin wild type and phosphovariants in cell lines devoid of endogenous L-plastin. The established cell lines were used for cell biology experiments and confocal microscopy analysis. Results Our modelling approach revealed that, in addition to the ERK/MAPK pathway and depending on the cellular context, the PI3K pathway contributes to L-plastin Ser5 phosphorylation through its downstream kinase SGK3. The results of the transwell invasion/migration assays showed that shRNA-mediated knockdown of L-plastin in BT-20 or HCC38 cells significantly reduced cell invasion, whereas stable expression of the phosphomimetic L-plastin Ser5Glu variant led to increased migration and invasion of BT-549 and MDA-MB-231 cells. Finally, confocal image analysis combined with zymography experiments and gelatin degradation assays provided evidence that L-plastin Ser5 phosphorylation promotes L-plastin recruitment to invadopodia, MMP-9 activity and concomitant extracellular matrix degradation. Conclusion Altogether, our results demonstrate that L-plastin Ser5 phosphorylation increases breast cancer cell invasiveness. Being a downstream molecule of both ERK/MAPK and PI3K/SGK pathways, L-plastin is proposed here as a potential target for therapeutic approaches that are aimed at blocking dysregulated signalling outcome of both pathways and, thus, at impairing cancer cell invasion and metastasis formation. Video abstracthttps://doi.org/10.1186/s12964-021-00710-5L-plastinActin-bundlingPI3K pathwayERK/MAPK pathwaySGKRSK |
| spellingShingle | Raquel A. C. Machado Dunja Stojevski Sébastien De Landtsheer Philippe Lucarelli Alexandre Baron Thomas Sauter Elisabeth Schaffner-Reckinger L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness Cell Communication and Signaling L-plastin Actin-bundling PI3K pathway ERK/MAPK pathway SGK RSK |
| title | L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness |
| title_full | L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness |
| title_fullStr | L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness |
| title_full_unstemmed | L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness |
| title_short | L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness |
| title_sort | l plastin ser5 phosphorylation is modulated by the pi3k sgk pathway and promotes breast cancer cell invasiveness |
| topic | L-plastin Actin-bundling PI3K pathway ERK/MAPK pathway SGK RSK |
| url | https://doi.org/10.1186/s12964-021-00710-5 |
| work_keys_str_mv | AT raquelacmachado lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness AT dunjastojevski lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness AT sebastiendelandtsheer lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness AT philippelucarelli lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness AT alexandrebaron lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness AT thomassauter lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness AT elisabethschaffnerreckinger lplastinser5phosphorylationismodulatedbythepi3ksgkpathwayandpromotesbreastcancercellinvasiveness |