Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma
Abstract Background Melanoma cells frequently dedifferentiate in response to inflammation which can increase responses to certain cytokines. Interferon-γ (IFNγ) is an integral part of the anti-tumor immune response and can directly induce both differentiational changes and expression of immunosuppre...
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BMC
2024-12-01
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| Series: | Cell Communication and Signaling |
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| Online Access: | https://doi.org/10.1186/s12964-024-01963-6 |
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| author | Teitur Sævarsson Adrián López García de Lomana Ólafur Sánchez Veerle van Esch Gunnar Bjarni Ragnarsson Siggeir Fannar Brynjólfsson Eiríkur Steingrímsson Berglind Ósk Einarsdóttir |
| author_facet | Teitur Sævarsson Adrián López García de Lomana Ólafur Sánchez Veerle van Esch Gunnar Bjarni Ragnarsson Siggeir Fannar Brynjólfsson Eiríkur Steingrímsson Berglind Ósk Einarsdóttir |
| author_sort | Teitur Sævarsson |
| collection | DOAJ |
| description | Abstract Background Melanoma cells frequently dedifferentiate in response to inflammation which can increase responses to certain cytokines. Interferon-γ (IFNγ) is an integral part of the anti-tumor immune response and can directly induce both differentiational changes and expression of immunosuppressive proteins in melanoma cells. How the differentiation status of melanoma cells affects IFNγ responses remains unclear. Methods Dedifferentiation of melanoma cells was induced via either siRNA or shRNA mediated MITF knockdown and the cells were subsequently treated with IFNγ. Effects of MITF knockdown and IFNγ treatment on gene expression were evaluated via qPCR and RNA sequencing. A Luminex assay was used to analyze the effects of dedifferentiation and IFNγ treatment on cytokine secretion. Effects on PD-L1 protein expression were analyzed via flow cytometry and western blotting. Inhibition of the JAK kinases, NF-κB and STAT3 with small molecule inhibitors, and siRNA mediated knockdown of STAT1 and IRF1 was applied to investigate the molecular mechanism behind IFNγ induced PD-L1 expression in dedifferentiated melanoma cells. The effects of inhibitor treatments and siRNA mediated knockdowns were evaluated via qPCR and western blotting. Bioinformatic analysis of publicly available RNA sequencing data, consisting of 45 patient derived melanoma cell lines, with or without IFNγ treatment, was conducted to assess the generalizability of the in vitro results. Results Dedifferentiation renders 624Mel melanoma cells hypersensitive to IFNγ stimulation in a context-dependent manner, resulting in non-additive upregulation of IFNγ-induced genes, increased PD-L1 protein expression and amplified secretion of CCL2, CXCL10 and IL-10. Furthermore, the intensified PD-L1 protein expression occurs through the JAK-STAT1-IRF1 axis. Lastly, dedifferentiated patient derived melanoma cell lines showed enhanced inflammatory signaling in response to IFNγ compared to differentiated cells, and tended to have higher PD-L1 expression, associated with increased IRF1 expression and activity. Conclusions Together, these findings indicate the existence of a molecular context linking dedifferentiation and IFNγ signaling in melanoma which may lead to immune evasion. Additionally, the variability in PD-L1 expression among MITFlow and MITFhigh cells suggests that high IFNγ-induced PD-L1 expression associates with enhanced inflammatory gene expression. These results imply that modulating melanoma differentiation may help shape IFNγ responsiveness. Graphical Abstract |
| format | Article |
| id | doaj-art-1d78e79fdd7343229e570bc587f79031 |
| institution | Kabale University |
| issn | 1478-811X |
| language | English |
| publishDate | 2024-12-01 |
| publisher | BMC |
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| series | Cell Communication and Signaling |
| spelling | doaj-art-1d78e79fdd7343229e570bc587f790312025-01-05T12:41:41ZengBMCCell Communication and Signaling1478-811X2024-12-0122112310.1186/s12964-024-01963-6Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanomaTeitur Sævarsson0Adrián López García de Lomana1Ólafur Sánchez2Veerle van Esch3Gunnar Bjarni Ragnarsson4Siggeir Fannar Brynjólfsson5Eiríkur Steingrímsson6Berglind Ósk Einarsdóttir7Department of Biomedical Science, Faculty of Medicine, BioMedical Center, University of IcelandDepartment of Biochemistry and Molecular Biology, Faculty of Medicine, BioMedical Center, University of IcelandDepartment of Biomedical Science, Faculty of Medicine, BioMedical Center, University of IcelandDepartment of Biomedical Science, Faculty of Medicine, BioMedical Center, University of IcelandLandspítali – The National University Hospital of IcelandDepartment of Immunology, Landspítali – The National University Hospital of IcelandDepartment of Biochemistry and Molecular Biology, Faculty of Medicine, BioMedical Center, University of IcelandDepartment of Biomedical Science, Faculty of Medicine, BioMedical Center, University of IcelandAbstract Background Melanoma cells frequently dedifferentiate in response to inflammation which can increase responses to certain cytokines. Interferon-γ (IFNγ) is an integral part of the anti-tumor immune response and can directly induce both differentiational changes and expression of immunosuppressive proteins in melanoma cells. How the differentiation status of melanoma cells affects IFNγ responses remains unclear. Methods Dedifferentiation of melanoma cells was induced via either siRNA or shRNA mediated MITF knockdown and the cells were subsequently treated with IFNγ. Effects of MITF knockdown and IFNγ treatment on gene expression were evaluated via qPCR and RNA sequencing. A Luminex assay was used to analyze the effects of dedifferentiation and IFNγ treatment on cytokine secretion. Effects on PD-L1 protein expression were analyzed via flow cytometry and western blotting. Inhibition of the JAK kinases, NF-κB and STAT3 with small molecule inhibitors, and siRNA mediated knockdown of STAT1 and IRF1 was applied to investigate the molecular mechanism behind IFNγ induced PD-L1 expression in dedifferentiated melanoma cells. The effects of inhibitor treatments and siRNA mediated knockdowns were evaluated via qPCR and western blotting. Bioinformatic analysis of publicly available RNA sequencing data, consisting of 45 patient derived melanoma cell lines, with or without IFNγ treatment, was conducted to assess the generalizability of the in vitro results. Results Dedifferentiation renders 624Mel melanoma cells hypersensitive to IFNγ stimulation in a context-dependent manner, resulting in non-additive upregulation of IFNγ-induced genes, increased PD-L1 protein expression and amplified secretion of CCL2, CXCL10 and IL-10. Furthermore, the intensified PD-L1 protein expression occurs through the JAK-STAT1-IRF1 axis. Lastly, dedifferentiated patient derived melanoma cell lines showed enhanced inflammatory signaling in response to IFNγ compared to differentiated cells, and tended to have higher PD-L1 expression, associated with increased IRF1 expression and activity. Conclusions Together, these findings indicate the existence of a molecular context linking dedifferentiation and IFNγ signaling in melanoma which may lead to immune evasion. Additionally, the variability in PD-L1 expression among MITFlow and MITFhigh cells suggests that high IFNγ-induced PD-L1 expression associates with enhanced inflammatory gene expression. These results imply that modulating melanoma differentiation may help shape IFNγ responsiveness. Graphical Abstracthttps://doi.org/10.1186/s12964-024-01963-6MelanomaDedifferentiationInterferon-γJAK/STAT pathwayPD-L1 |
| spellingShingle | Teitur Sævarsson Adrián López García de Lomana Ólafur Sánchez Veerle van Esch Gunnar Bjarni Ragnarsson Siggeir Fannar Brynjólfsson Eiríkur Steingrímsson Berglind Ósk Einarsdóttir Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma Cell Communication and Signaling Melanoma Dedifferentiation Interferon-γ JAK/STAT pathway PD-L1 |
| title | Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma |
| title_full | Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma |
| title_fullStr | Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma |
| title_full_unstemmed | Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma |
| title_short | Differentiation status determines the effects of IFNγ on the expression of PD-L1 and immunomodulatory genes in melanoma |
| title_sort | differentiation status determines the effects of ifnγ on the expression of pd l1 and immunomodulatory genes in melanoma |
| topic | Melanoma Dedifferentiation Interferon-γ JAK/STAT pathway PD-L1 |
| url | https://doi.org/10.1186/s12964-024-01963-6 |
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