Neuronal Dot1l Activity Acts as a Mitochondrial Gene-Repressor Associated with Human Brain Aging via H3K79 Hypermethylation
Abstract
:1. Introduction
2. Results
2.1. Neuronal Dot1l Transcript Abundance Increases through H2O2 and AMPK Activation
2.2. Fast Turnover of H3K79 Methylation in Post-Mitotic DA Neurons
2.3. Conditional Dot1l Deletion Leads to a Rostro-Lateral Non-Progressive Defect in Levels of Dopaminergic Markers
2.4. Dot1l Ablation Leads to Increased Mitochondrial Respiratory Chain Transcripts
2.5. Respiratory Chain Transcript Abundance Remains up in 6 Months Old Neurons with Reduced Dot1l
2.6. mtDNA Encoded Genes and a Selection of Genes Involved in Mitochondrial Morphology Form Exceptions
3. Discussion
3.1. The Relation between Dot1l and H3K79me2 Deregulation and Cellular Stress
3.2. Influencing Dot1l Activity to Alleviate Aging Mechanisms and Neuronal Pathology
3.3. Transcriptional Exceptions: mtDNA Encoded Respiratory Genes
3.4. Regulation of Mitochondrial Turnover and Mitogenesis
4. Methods and Materials
4.1. Human Tissue and Anatomical Selection
4.2. Immunohistochemistry Human Brain Tissue
4.3. Imaging
4.4. Quantification of IMHC Signal Humane Tissue
4.5. Animals
4.6. Immunohistochemistry Mouse Brain Tissue
4.7. Primary Neuronal Culturing and Treatment
4.8. In Situ Hybridization
Antibody | Product Number | Concentration |
rabbit-α-H3K79me2 | Abcam, Ab3594 | 1:1000 in THZT |
rabbit-α-H3K79me3 | Abcam, Ab2621 | 1:750 in THZT |
Rabbit-α-H3K27me3 | Millipore, 17–622 | 1:1000 in THZT |
sheep-α-TH | Millipore, Ab1542 | 1:500 in THZT |
Goat-α-rabbit-555 | Life Technologies, A27039 | 1:1000 in PBS |
donkey-α-sheep-488 | Life Technologies, A11015 | 1:500 in TBS |
chicken-α-GFP | Chicken-anti-GFP | 1:1000 in THZT |
rabbit-α-H3K79me1 | Abcam, ab2886 | 1:100 in THZT |
Goat-α-chicken-488 | Thermo Fisher, A-11039 | 1:1000 in PBS |
Donkey-α-rabbit-594 | Thermo Fisher, A-21207 | 1:1000 in PBS |
4.9. Fluorescence Associated Cell Sorting (FACS)
4.10. RNA Sample Generation
4.11. RNA Sequencing
4.12. Quantitative PCR
Target | Forward Primer | Reversed Primer |
Th | TGCACACAGTACATCCGTCATGC | GCAAATGTGCGGTACGCCAACA |
Vmat2 | CCTCTTACGACCTTGCTGAAGG | GCTGCCACTTTCGGGAACACAT |
Dat | GGTGCTGATTGCCTTCTCCAGT | GACAACGAAGCCAGAGGAGAAG |
Pitx3 | CCTTCCAGAGGAATCGCTACCT | CTGCGAAGCCACCTTGCACA |
Rspo2 | CCGAGCCCCAGATATGAACA | GACCAACTTCACAACCTTCTACA |
Calb2 | TGACTGCATCCCAGTTCCTG | CTTGGACATCATGCCAGAACC |
Cck | TAGCGCGATACATCCAGCAGGT | GGTATTCGTAGTCCTCGGCACT |
Girk2 | GGAACTGGAGATTGTGGTCATCC | TCTTCCAGCGTTAGGACAGCTTCCAGAGTG |
Ahd2 | GGAATACCGTGGTTGTCAAGCC | CCAGGGACAATGTTTACCACGC |
Nurr1 | CCGCCGAAATCGTTGTCAGTAC | TTCGGCTTCGAGGGTAAACGAC |
TBP | GAGAATAAGAGAGCCACGC | TCACATCACAGCTCCCCAC |
Dot1l | CTGGTGGCCCAGATGATTGA | GTCCATGGTCTCTGCGTACT |
4.13. Statistics and Graphs
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Van Heesbeen, H.J.; Von Oerthel, L.; De Vries, P.M.; Wagemans, C.M.R.J.; Smidt, M.P. Neuronal Dot1l Activity Acts as a Mitochondrial Gene-Repressor Associated with Human Brain Aging via H3K79 Hypermethylation. Int. J. Mol. Sci. 2023, 24, 1387. https://doi.org/10.3390/ijms24021387
Van Heesbeen HJ, Von Oerthel L, De Vries PM, Wagemans CMRJ, Smidt MP. Neuronal Dot1l Activity Acts as a Mitochondrial Gene-Repressor Associated with Human Brain Aging via H3K79 Hypermethylation. International Journal of Molecular Sciences. 2023; 24(2):1387. https://doi.org/10.3390/ijms24021387
Chicago/Turabian StyleVan Heesbeen, Hendrikus J., Lars Von Oerthel, Paul M. De Vries, Cindy M. R. J. Wagemans, and Marten P. Smidt. 2023. "Neuronal Dot1l Activity Acts as a Mitochondrial Gene-Repressor Associated with Human Brain Aging via H3K79 Hypermethylation" International Journal of Molecular Sciences 24, no. 2: 1387. https://doi.org/10.3390/ijms24021387
APA StyleVan Heesbeen, H. J., Von Oerthel, L., De Vries, P. M., Wagemans, C. M. R. J., & Smidt, M. P. (2023). Neuronal Dot1l Activity Acts as a Mitochondrial Gene-Repressor Associated with Human Brain Aging via H3K79 Hypermethylation. International Journal of Molecular Sciences, 24(2), 1387. https://doi.org/10.3390/ijms24021387