YB-1 Phosphorylation at Serine 209 Inhibits Its Nuclear Translocation
Abstract
:1. Introduction
2. Results
2.1. Serum Stimulation Does Not Induce YB-1 Nuclear Translocation in HeLa, NIH3T3, and PC-3 Cells
2.2. Akt Kinase Phosphorylates YB-1 at the S209 Residue In Vitro
2.3. S209 Phosphorylation Inhibits YB-1 Nuclear Translocation
2.4. S209 Phosphorylation Does Not Change the RNA-Binding Ability of YB-1
2.5. The Effect of S209 Phosphorylation on YB-1 Transport Prevails over That of S102 Phosphorylation
3. Discussion
4. Materials and Methods
4.1. Expression Constructs and Recombinant Proteins
4.2. In Vitro Kinase Assay
4.3. Electrophoretic Mobility Shift Assay (EMSA)
4.4. Cell Cultivation and Transfection
4.5. Western Blot
4.6. In Vitro Transport Assay
4.7. Immunofluorescence Microscopy (IF)
4.8. Phosphorylation Site Predictions and Conservation Analysis
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Acknowledgments
Conflicts of Interest
References
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Primer name | Sequence | Comments |
---|---|---|
For T80A | 5′-CATCTTCCTTGGCGTCATTCCTG-3′ | 1st step PCR forward primers |
For T80D | 5′-CATCTTCCTTGACGTCATTCCTG-3′ | |
For S102A | 5′-GTACCTTCGCGCTGTAGGAGATGGAG-3′ | |
For S102D | 5′-GTACCTTCGCGACGTAGGAGATGGAG-3′ | |
For S209A | 5′-CCACAGTATGCTAACCCTCCTG-3′ | |
For S209D | 5′-CCACAGTATGACAACCCTCCTG-3′ | |
For_NdeI | 5′-ATATACATATGAGCAGCGAGGCCGAGAC-3′ | 2nd step PCR forward primer, NdeI site is underlined |
Rev_BamHI | 5′-TCTTCGGATCCAATCTTTTGTTCATTTC-3′ | 1st and 2nd steps PCR reverse primer, BamHI site is underlined |
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Sogorina, E.M.; Kim, E.R.; Sorokin, A.V.; Lyabin, D.N.; Ovchinnikov, L.P.; Mordovkina, D.A.; Eliseeva, I.A. YB-1 Phosphorylation at Serine 209 Inhibits Its Nuclear Translocation. Int. J. Mol. Sci. 2022, 23, 428. https://doi.org/10.3390/ijms23010428
Sogorina EM, Kim ER, Sorokin AV, Lyabin DN, Ovchinnikov LP, Mordovkina DA, Eliseeva IA. YB-1 Phosphorylation at Serine 209 Inhibits Its Nuclear Translocation. International Journal of Molecular Sciences. 2022; 23(1):428. https://doi.org/10.3390/ijms23010428
Chicago/Turabian StyleSogorina, Ekaterina M., Ekaterina R. Kim, Alexey V. Sorokin, Dmitry N. Lyabin, Lev P. Ovchinnikov, Daria A. Mordovkina, and Irina A. Eliseeva. 2022. "YB-1 Phosphorylation at Serine 209 Inhibits Its Nuclear Translocation" International Journal of Molecular Sciences 23, no. 1: 428. https://doi.org/10.3390/ijms23010428
APA StyleSogorina, E. M., Kim, E. R., Sorokin, A. V., Lyabin, D. N., Ovchinnikov, L. P., Mordovkina, D. A., & Eliseeva, I. A. (2022). YB-1 Phosphorylation at Serine 209 Inhibits Its Nuclear Translocation. International Journal of Molecular Sciences, 23(1), 428. https://doi.org/10.3390/ijms23010428