The Ash2l SDI Domain Is Required to Maintain the Stability and Binding of DPY30
Abstract
:1. Introduction
2. Materials and Methods
2.1. Animals
2.2. Cell Culture
2.3. Tissue Section and Immunofluorescence
2.4. Plasmid Construction
2.5. RT-qPCR
2.6. Western Blot and Coimmunoprecipitation
2.7. RNA-Seq and ChIP-Seq Analysis
3. Results
3.1. ASH2L Is Necessary for the Expression of DPY30
3.2. The Stability of DPY30 Relies on Direct Binding with ASH2L
3.3. The Degradation of DPY30 Relies on the Ubiquitin-Proteasome System
3.4. DPY30 Is Capable of Ameliorating the Outcome of ASH2L Depletion
4. Discussion
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Name | Sequence |
---|---|
hAsh2l-F | ATGGATACTCAGGCGGGCTC |
hAsh2l-R | TCAGGGTTCCCATGGGGG |
hAsh2l-ΔSDI-F | ATGGATACTCAGGCGGGCTC |
hAsh2l-ΔSDI-R | CATGTCACTCATAGGGCGGTAA |
hAsh2l-3E-F | ACCCTTGCTGACGTCCTCTATCACCTCGAGACAGAAGTGGATGGGAGGCG |
hAsh2l-3E-R | CTCGAGGTGATAGAGGACGTCAGCAAGGGTGTGCTCTACCACGGCGCC |
Ash2l-qPCR-F | CCTACTTTCTCCGGAAGCAAG |
Ash2l-qPCR-R | GACAATGTTATTGGGCCAAGTC |
Dpy30-qPCR-F | AACGCAGGTTGCAGA AAATCCT |
Dpy30-qPCR-R | TCTGATCCAGGTAGGCACGAG |
Ccnd1-qPCR-F | CTGATTGGACAGGCATGGGT |
Ccnd1-qPCR-R | GTGCCTGGAAGTCAACGGTA |
Gapdh-qPCR-F | GGTCATCCATGACAACTTTGG |
Gapdh-qPCR-R | GGCCATCACGCCACAG |
siAsh2l-1# | CAGTAAAGATCCAGAAGAA |
siAsh2l-2# | GGTGCCTGGTATTTTGAAA |
siAsh2l-3# | CGAAGACAATGTTCTCCAA |
mAsh2l-shRNA-1 | CAAGAAGGCCAGAAGTGATCCTTTATTCAAGAGATAAAGGATCACTTCTGGCCTTCTTGTTTTTT |
mAsh2l-shRNA-3 | CCTGTGTCTGTGTGTTCCAAATTCAAGAGATTTGGAACACACAGACACAGGTTTTTT |
hAsh2l-shRNA-1 | CCGGGTGACTTGTTATCCTACTATACTCGAGTATAGTAGGATAACAAGTCACTTTTT |
hAsh2l-shRNA-2 | CCGGCCGAAGACAATGTTCTCCAAACTCGAGTTTGGAGAACATTGTCTTCGGTTTTT |
hAsh2l-shRNA-3 | CCGGCCTGCTTGTATGAACGGGTTTCTCGAGAAACCCGTTCATACAAGCAGGTTTTT |
Name | Resource |
---|---|
Anti-Ash2 | BETHYL (Waltham, MA, USA, #A300–112A) |
Anti-DPY30 | Sigma (St. Louis, MO, USA, #HPA043761) |
Anti-WDR5 | R&D system (Minneapolis, MN, USA, #AF5810) |
Anti-RbBP5 | Abcam (Cambridge, UK, #ab52084) |
Anti-Cyclin D1 | Abcam (Cambridge, UK, #ab16663) |
Anti-FLAG | ABclonal (Cambridge, UK, #WH166500) |
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Ma, M.; Zhou, J.; Ma, Z.; Chen, H.; Li, L.; Hou, L.; Yin, B.; Qiang, B.; Shu, P.; Peng, X. The Ash2l SDI Domain Is Required to Maintain the Stability and Binding of DPY30. Cells 2022, 11, 1450. https://doi.org/10.3390/cells11091450
Ma M, Zhou J, Ma Z, Chen H, Li L, Hou L, Yin B, Qiang B, Shu P, Peng X. The Ash2l SDI Domain Is Required to Maintain the Stability and Binding of DPY30. Cells. 2022; 11(9):1450. https://doi.org/10.3390/cells11091450
Chicago/Turabian StyleMa, Mengjie, Jiafeng Zhou, Zhihua Ma, Hanxue Chen, Liang Li, Lin Hou, Bin Yin, Boqin Qiang, Pengcheng Shu, and Xiaozhong Peng. 2022. "The Ash2l SDI Domain Is Required to Maintain the Stability and Binding of DPY30" Cells 11, no. 9: 1450. https://doi.org/10.3390/cells11091450