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The effect of phosphoserine aminotransferase-1 dependent fluctuation on timing of embryonic stem cell differentiation
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | 윤홍덕 | - |
| dc.contributor.author | 황인영 | - |
| dc.date.accessioned | 2017-07-14T01:46:05Z | - |
| dc.date.available | 2017-07-14T01:46:05Z | - |
| dc.date.issued | 2016-08 | - |
| dc.identifier.other | 000000136369 | - |
| dc.identifier.uri | https://hdl.handle.net/10371/122324 | - |
| dc.description | 학위논문 (박사)-- 서울대학교 대학원 : 의과학과, 2016. 8. 윤홍덕. | - |
| dc.description.abstract | Embryonic stem cells (ESCs) undergo coordinated epigenetic and metabolic changes to differentiate properly. However, the precise mechanisms by which these alterations are finely tuned at early stages of differentiation have not been identified. In this study, we demonstrate that phosphoserine aminotransferase 1 (Psat1), an Oct4/Sox2/Nanog target protein that regulates changes in α-ketoglutarate (α-KG), determines the fate of mESCs. Maintaining Psat1 level was essential for mESCs self-renewal and pluripotency. Moderate Psat1 knockdown mESCs had decreased DNA 5-hydroxymethylation (5-hmC) and increased histone methylation levels by downregulating permissive amounts of α-KG, ultimately accelerating differentiation. We found that intracellular α-KG transiently declined during differentiation and its disturbance by treatment with dimethyl-α-KG (dm-α-KG) delayed differentiation. Further, in vivo teratoma formation assay showed impaired pluripotency of Psat1 knockdown mESCs especially into ectodermal lineage. Altogether, we reveal how Psat1 is regulated to maintain intracellular α-KG and the fate of mESCs | - |
| dc.description.tableofcontents | I. INTRODUCTION 1
1-1. Master transcription factors in mESCs 2 1-2. Epigenetic changes during ESC differentiation 3 1-3. Metabolism and epigenetic modifications 3 1-4. Metabolic features of ESCs 3 II. MATERIALS AND METHODS 9 2-1. Stable isotope labeling 10 2-2. LC-MS 10 2-3. Metabolite extraction for LC-MS 11 2-4. Histone purification 11 2-5. Hydroxymethylated DNA immunoprecipitation 12 2-6. Dot blot assay 12 2-7. In vivo teratoma formation assay 12 2-8. Histological analysis 13 2-9. Western blot analysis 13 2-10. shRNA mediated knockdown 13 2-11. Stable cell line generation 13 2-12. AP staining 14 2-13. Genomic DNA preparation 14 2-14. ChIP 14 2-15. Quantitative real time PCR 14 2-16. Cell culture 15 2-17. Statistical analysis 15 III. RESULTS 20 3-1. Psat1 is Regulated by Core Transcription Factors in mESCs 21 3-2. Psat1 Regulates Intracellular α-KG Levels in mESCs 37 3-3. Psat1 Regulates Histone and DNA Methylation States 49 3-4. α-KG Levels Affect the Timing of mESC Differentiation 62 IV. DISCUSSION 76 V. REFERENCES 81 Ⅵ. ABSTRACT IN KOREAN 90 | - |
| dc.format | application/pdf | - |
| dc.format.extent | 7764047 bytes | - |
| dc.format.medium | application/pdf | - |
| dc.language.iso | en | - |
| dc.publisher | 서울대학교 대학원 | - |
| dc.subject | Stem cell | - |
| dc.subject | Metabolism | - |
| dc.subject | Epigenetics | - |
| dc.subject | Psat-1 α-ketoglutarate | - |
| dc.subject.ddc | 610 | - |
| dc.title | The effect of phosphoserine aminotransferase-1 dependent fluctuation on timing of embryonic stem cell differentiation | - |
| dc.type | Thesis | - |
| dc.description.degree | Doctor | - |
| dc.citation.pages | 92 | - |
| dc.contributor.affiliation | 의과대학 의과학과 | - |
| dc.date.awarded | 2016-08 | - |
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