Characterization of HLA-G Regulation and HLA Expression in Breast Cancer and Malignant Melanoma Cell Lines upon IFN-γ Stimulation and Inhibition of DNA Methylation
Abstract
:1. Introduction
2. Results
2.1. HLA Cell Surface Expression in Breast Cancer and Malignant Melanoma Cell Lines upon IFN-γ Stimulation
2.2. HLA Cell Surface Expression in Breast Cancer and Malignant Melanoma Cell Lines upon Demethylation Treatment
2.3. HLA-G mRNA Expression in Breast Cancer and Malignant Melanoma Cell Lines
2.4. Verification of HLA-G Demethylation
2.5. Detection of HLA-G mRNA Isoforms in Breast Cancer and Malignant Melanoma Cell Lines upon 5-aza-dC Treatment
3. Discussion
4. Materials and Methods
4.1. Cells and Culture Conditions
4.2. Cell Treatments
4.3. Flow Cytometry
4.4. DNA and RNA Extraction and cDNA Synthesis
4.5. Digital Droplet PCR
4.6. PCR and Fragment Analysis
4.7. Genotyping of the HLA-G 14 bp Ins/Del Polymorphism
4.8. Bisulfite Sequencing
4.9. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Cell Line | 14 bp Polymorphism |
---|---|
MDA-MB-231 | del/del |
MCF-7 | del/del |
FM-55M2 | ins/del |
FM-56 | ins/ins |
JEG-3 | ins/ins |
Cell Line | Condition | HLA-A | HLA-B | HLA-C | HLA-E | HLA-G * | HLA-DR |
---|---|---|---|---|---|---|---|
MDA-MB-231 | Untreated ** | + | + | (+) *** | (+) | − | − |
IFN-γ | ↑ | ↑ | ↑ | ↑ | − | ↑ | |
5-Aza-dC | |||||||
10 µM, day 3 | ↑ | ↑ | − | − | ↑ | ↑ | |
10 µM, day 6 | ↑ | ↑ | ↑ | ↑ | - | ↑ | |
100 µM, day 3 | ↑ | ↑ | ↑ | ↓ | ↑ | ↑ | |
100 µM, day 6 | − | ↑ | ↑ | ↑ | − | ↑ | |
MCF-7 | Untreated | + | − | − | (+) | − | − |
IFN-γ | ↑ | ↑ | − | − | − | − | |
5-Aza-dC | |||||||
10 µM, day 3 | ↑ | ↑ | ↑ | - | ↑ | − | |
10 µM, day 6 | ↑ | ↑ | ↑ | ↑ | ↑ | − | |
100 µM, day 3 | − | − | − | − | ↑ | − | |
100 µM, day 6 | ↑ | ↑ | ↑ | - | ↑ | - | |
FM-55M2 | Untreated | + | + | − | (+) | − | (+) |
IFN-γ | ↑ | ↑ | ↑ | ↑ | − | ↑ | |
5-Aza-dC | |||||||
100 µM, day 3 | ↓ | ↑ | - | - | ↑ | − | |
100 µM, day 6 | ↓ | − | ↓ | ↑ | − | ↑ | |
FM-56 | Untreated | − | (+) | − | − | − | + |
IFN-γ | − | ↑ | ↑ | ↑ | − | ↑ | |
5-Aza-dC | |||||||
100 µM, day 3 | − | ↑ | − | − | ↑ | − | |
100 µM, day 6 | ↑ | ↑ | ↑ | ↑ | ↑ | − | |
JEG-3 | Untreated | (+) | (+) | + | + | + | − |
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Jørgensen, N.; Sayed, A.; Jeppesen, H.B.; Persson, G.; Weisdorf, I.; Funck, T.; Hviid, T.V.F. Characterization of HLA-G Regulation and HLA Expression in Breast Cancer and Malignant Melanoma Cell Lines upon IFN-γ Stimulation and Inhibition of DNA Methylation. Int. J. Mol. Sci. 2020, 21, 4307. https://doi.org/10.3390/ijms21124307
Jørgensen N, Sayed A, Jeppesen HB, Persson G, Weisdorf I, Funck T, Hviid TVF. Characterization of HLA-G Regulation and HLA Expression in Breast Cancer and Malignant Melanoma Cell Lines upon IFN-γ Stimulation and Inhibition of DNA Methylation. International Journal of Molecular Sciences. 2020; 21(12):4307. https://doi.org/10.3390/ijms21124307
Chicago/Turabian StyleJørgensen, Nanna, Abid Sayed, Helene Bjerregaard Jeppesen, Gry Persson, Iben Weisdorf, Tina Funck, and Thomas Vauvert Faurschou Hviid. 2020. "Characterization of HLA-G Regulation and HLA Expression in Breast Cancer and Malignant Melanoma Cell Lines upon IFN-γ Stimulation and Inhibition of DNA Methylation" International Journal of Molecular Sciences 21, no. 12: 4307. https://doi.org/10.3390/ijms21124307