Spatial oxidation of L-plastin downmodulates actin-based functions of tumor cells

Balta, Emre; Hardt, Robert; Liang, Jie; Kirchgessner, h.c. mult. M.; Orlik, Christian; Jahraus, Beate; Hillmer, Stefan; Meuer, Stefan; Hübner, Katrin; Wabnitz, Guido H.; Samstag, Yvonne

doi:10.1038/s41467-019-11909-z

Cited by 26 publications

(31 citation statements)

References 61 publications

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“…In addition, the podosomes merging from the sealing belt of large multinucleated cells also mediate osteoclast fusion (11). LPL regulates filopodia formation, and the inhibition of LPL by specific nanobodies or oxidation block filopodia formation (4,14). Here, we showed that filopodia initiated preosteoclast fusion and that LPL deletion inhibited filopodia formation and thus preosteoclast fusion.…”

Section: Discussionmentioning

confidence: 75%

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Targeting actin-bundling protein L-plastin as an anabolic therapy for bone loss

et al. 2020

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The actin-bundling protein L-plastin (LPL) mediates the resorption activity of osteoclasts, but its therapeutic potential in pathological bone loss remains unexplored. Here, we report that LPL knockout mice show increased bone mass and cortical thickness with more mononuclear tartrate-resistant acid phosphatase–positive cells, osteoblasts, CD31hiEmcnhi endothelial vessels, and fewer multinuclear osteoclasts in the bone marrow and periosteum. LPL deletion impeded preosteoclasts fusion by inhibiting filopodia formation and increased the number of preosteoclasts, which release platelet-derived growth factor-BB to promote CD31hiEmcnhi vessel growth and bone formation. LPL expression is regulated by the phosphatidylinositol 3-kinase/AKT/specific protein 1 axis in response to receptor activator of nuclear factor–κB ligand. Furthermore, we identified an LPL inhibitor, oroxylin A, that could maintain bone mass in ovariectomy-induced osteoporosis and accelerate bone fracture healing in mice. In conclusion, we showed that LPL regulates osteoclasts fusion, and targeting LPL serves as a novel anabolic therapy for pathological bone loss.

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Section: Discussionmentioning

confidence: 75%

“…LPL mediates filopodia formation in renal podocytes and cooperates with other bundling proteins to form filopodia in cancer cells (3,13). The inhibition of LPL by nanobody or oxidation diminishes the actin-bundling activity of LPL and leads to fewer filopodial extensions in cancer cells (4,14).…”

Section: Introductionmentioning

confidence: 99%

Targeting actin-bundling protein L-plastin as an anabolic therapy for bone loss

et al. 2020

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“…In a next step, we examined the functional impact of the L-plastin Ser5 phosphorylation event in breast cancer cells. Whereas calcium binding and oxidation of L-plastin inhibit the actin-bundling activity of L-plastin [ 60 – 63 ], Ser5 phosphorylation is known to promote its targeting to actin-rich structures and to increase its F-actin bundling activity in vitro and in cells [ 11 , 12 ]. L-plastin phosphorylation on residues Ser5 and Ser7 has been linked to bone resorption activity via nascent sealing zone and sealing ring formation in cultured osteoclasts and in mice [ 64 , 65 ].…”

Section: Discussionmentioning

confidence: 99%

L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness

et al. 2021

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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.

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“…Again, the bundling-inhibitor Nb5 had no effect on MMP-9 secretion and activity, con rming that Nb5-mediated reduction of degradation is not dependent on MMP-9 secretion and activity. More recently, Balta and collaborators highlighted the existence of a link between L-plastin expression, total MMP activity and MMP-2 release in MV3 melanoma cells (47). L-plastin and MMP-2 were found to localize in invadopodial extensions and to co-immunoprecipitate, indicating that Lplastin may help MMP-2 translocate to invadopodial structures.…”

Section: Discussionmentioning

confidence: 99%

“…In a next step, we examined the functional impact of the L-plastin Ser5 phosphorylation event in breast cancer cells. Whereas calcium binding and oxidation of L-plastin inhibit the actin-bundling activity of Lplastin (44)(45)(46)(47), Ser5 phosphorylation is known to promote its targeting to actin-rich structures and to increase its F-actin bundling activity in vitro and in cells (11,12). L-plastin phosphorylation on residues Ser5 and Ser7 has been linked to bone resorption activity via nascent sealing zone and sealing ring formation in cultured osteoclasts and in mice (48,49).…”

Section: Discussionmentioning

confidence: 99%

L-Plastin Ser5 Phosphorylation is Modulated by the PI3K/SGK Pathway and Promotes Breast Cancer Cell Invasiveness

Machado

Stojevski

Landtsheer

et al. 2020

Preprint

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BackgroundMetastasis 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. MethodsTo 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.ResultsOur 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. ConclusionAltogether, 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.

show abstract

Spatial oxidation of L-plastin downmodulates actin-based functions of tumor cells

Cited by 26 publications

References 61 publications

Targeting actin-bundling protein L-plastin as an anabolic therapy for bone loss

Targeting actin-bundling protein L-plastin as an anabolic therapy for bone loss

L-plastin Ser5 phosphorylation is modulated by the PI3K/SGK pathway and promotes breast cancer cell invasiveness

L-Plastin Ser5 Phosphorylation is Modulated by the PI3K/SGK Pathway and Promotes Breast Cancer Cell Invasiveness

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