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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 吳益群(Yi-Chun Wu) | |
| dc.contributor.author | Cheng-Yi Chen | en |
| dc.contributor.author | 陳成奕 | zh_TW |
| dc.date.accessioned | 2021-07-11T14:49:20Z | - |
| dc.date.available | 2025-08-31 | |
| dc.date.copyright | 2020-09-23 | |
| dc.date.issued | 2020 | |
| dc.date.submitted | 2020-08-31 | |
| dc.identifier.citation | 1. Garavito, M.F., H.Y. Narvaez-Ortiz, and B.H. Zimmermann, Pyrimidine Metabolism: Dynamic and Versatile Pathways in Pathogens and Cellular Development. J Genet Genomics, 2015. 42(5): p. 195-205.
2. Jones, M.E., Pyrimidine Nucleotide Biosynthesis in Animals: Genes, Enzymes, and Regulation of UMP Biosynthesis. Annu. Rev. Biochem, 1980. 49(1): p. 253-279. 3. Mainguet, S.E., et al., Uracil salvage is necessary for early Arabidopsis development. Plant J, 2009. 60(2): p. 280-91. 4. Ng, S.B., et al., Exome sequencing identifies the cause of a mendelian disorder. Nat Genet, 2010. 42(1): p. 30-5. 5. Bailey, C.J., Orotic aciduria and uridine monophosphate synthase: a reappraisal. J Inherit Metab Dis, 2009. 32 Suppl 1: p. S227-33. 6. Webster, D.R., et al., Hereditary Orotic Aciduria and Other Disorders of Pyrimidine Metabolism, in The Online Metabolic and Molecular Bases of Inherited Disease, D.L. Valle, et al., Editors. 2019, McGraw-Hill Education: New York, NY. 7. Traut, T.W. and M.E. Jones, Uracil Metabolism—UMP Synthesis from Orotic Acid or Uridine and Conversion of Uracil to β-Alanine: Enzymes and cDNAs. 1996. p. 1-78. 8. Balasubramaniam, S., J.A. Duley, and J. Christodoulou, Inborn errors of pyrimidine metabolism: clinical update and therapy. J. Inherit. Metab. Dis., 2014. 37(5): p. 687-698. 9. Elmore, S., Apoptosis: a review of programmed cell death. Toxicol Pathol, 2007. 35(4): p. 495-516. 10. Kerr, J.F.R., C.M. Winterford, and B.V. Harmon, Apoptosis. Its significance in cancer and cancer Therapy. Cancer, 1994. 73(8): p. 2013-2026. 11. Wong, R.S., Apoptosis in cancer: from pathogenesis to treatment. J Exp Clin Cancer Res, 2011. 30(1): p. 87. 12. Sulston, J.E. and H.R. Horvitz, Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev. Biol., 1977. 56(1): p. 110-156. 13. Conradt, B., Y.C. Wu, and D. Xue, Programmed Cell Death During Caenorhabditis elegans Development. Genetics, 2016. 203(4): p. 1533-62. 14. Horvitz, H.R., Worms, Life, and Death (Nobel Lecture). ChemBioChem, 2003. 4(8): p. 697-711. 15. Hatzold, J. and B. Conradt, Control of apoptosis by asymmetric cell division. PLoS Biol, 2008. 6(4): p. e84. 16. Teuliere, J., et al., Asymmetric Neuroblast Divisions Producing Apoptotic Cells Require the Cytohesin GRP-1 in lt;em gt;Caenorhabditis elegans lt;/em gt. Genetics, 2014. 198(1): p. 229. 17. Huang, M., et al., Caspase-dependent cleavage of carbamoyl phosphate synthetase II during apoptosis. Mol Pharmacol, 2002. 61(3): p. 569-77. 18. Longley, D.B., D.P. Harkin, and P.G. Johnston, 5-Fluorouracil: mechanisms of action and clinical strategies. Nat. Rev. Cancer, 2003. 3(5): p. 330-338. 19. Zhang, C. and M. Chu, Leflunomide: A promising drug with good antitumor potential. BIOCHEM BIOPH RES CO, 2018. 496(2): p. 726-730. 20. Ren, A., et al., Leflunomide inhibits proliferation and tumorigenesis of oral squamous cell carcinoma. Mol Med Rep, 2017. 16(6): p. 9125-9130. 21. Fire, A., et al., Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature, 1998. 391(6669): p. 806-11. 22. Dokshin, G.A., et al., Robust Genome Editing with Short Single-Stranded and Long, Partially Single-Stranded DNA Donors in Caenorhabditis elegans. Genetics, 2018. 210(3): p. 781-787. 23. Xie, C.Y., et al., Dietary supplement with nucleotides in the form of uridine monophosphate or uridine stimulate intestinal development and promote nucleotide transport in weaned piglets. J Sci Food Agric, 2019. 99(13): p. 6108-6113. 24. Blackwell, T.K., et al., TOR Signaling in lt;em gt;Caenorhabditis elegans Development, Metabolism, and Aging. Genetics, 2019. 213(2): p. 329. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/78283 | - |
| dc.description.abstract | 計畫性細胞凋亡在生物的發育上扮演著重要的角色。在線蟲當中，主要有四個核心基因執行細胞凋亡，分別為: egl-1, ced-9, ced-4 以及 ced-3。 線蟲細胞凋亡的基因與人類的細胞凋亡基因具有高度保留性，到目前為止，我們對調控細胞凋亡仍所知有限，例如特定的細胞是否有不同的基因調控其凋亡。 在我們實驗室先前的研究中發現了，在不對稱分裂基因 grp-1 的突變株中，如果細胞凋亡核心基因同時發生變異，則會高機率在尾部產生多餘的表皮細胞 hyp8/9，並且會導致線蟲的尾部產生球狀突起，我們把這個性狀稱之為:「突出尾巴」 (bulged tail)。在grp-1 突變株的背景下，我們進行了基因篩選，來找出新的可以調控細胞凋亡的基因。我們發現新的可以調控細胞凋亡的基因:pyr-1，也就是人類的 CAD (carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase) 同源基因，PYR-1 為一嘧啶合成酶，負責催化嘧啶生合成途徑的前三個步驟，而嘧啶在生物體內中參與了許多重要的生理反應，例如:DNA 或RNA 的合成；磷脂質及尿苷二磷酸葡萄糖(UDP-sugar)的合成。在此篇研究中，我們進一步證明了在嘧啶合成途徑中，pyr-1 下游基因dhod-1 和 umps-1 同樣是進行細胞凋亡的必要基因，然而，在umps-1之後的嘧啶合成基因並不是細胞凋亡所需要的基因。此外，藉由額外的嘧啶補充劑及調控其在線蟲體內的代謝，我們發現UMP含量是造成hyp8/9細胞凋亡的成因。 更重要的是，在人類的子宮頸癌細胞株HeLa中抑制CAD 的表現同樣的保護了HeLa 細胞免於紫外線照射誘發的細胞凋亡。 總結來說，我們的實驗結果證實了 UMP可以引起細胞凋亡並且指出UMP引起細胞凋亡的能力可能在不同生物間是共有的。 | zh_TW |
| dc.description.abstract | Programmed cell death (PCD) is important for animal development. In Caenorhabditis elegans (C. elegans), the execution of PCD is controlled by four genes, egl-1, ced-9, ced-4, and ced-3, and this core PCD pathway is highly conserved from nematodes to humans. Despite of extensive studies in PCD, how different cells may be differentially regulated to undergo PCDs is not yet clear. Previous studies in our lab found that mutations in the core PCD pathway caused extra hypodermal hyp8/9 cells in a sensitized mutant background and resulted in a bulged tail phenotype. In a sensitized genetic screen, a pyr-1 mutant was isolated with a bulged tail. pyr-1 is an ortholog of human CAD (carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase) that catalyzes the first three steps of the de novo pyrimidine synthesis pathway. This pyrimidine synthesis pathway is important for DNA and RNA synthesis as well as the formation of phospholipids and UDP-sugars. In this thesis, we show that like pyr-1, dhod-1 and umps-1, two genes acting downstream of pyr-1 in the pyrimidine synthesis pathway, are required for normal PCD. Using genetic manipulation and metabolite supplementation, we show that the appropriate level of UMP, the end product produced by DHOD-1 and UMPS-1, is necessary for the cells to undergo PCD normally in the hyp8/9 lineage. Similar to the pro-apoptotic activity of pyr-1, knockdown of CAD in HeLa cells by RNA interference protects the cells from death under the UV treatment. These data show an unexpected role of UMP in regulation of programmed cell death and suggest evolutionary conservation of the pro-apoptotic function of UMP in C. elegans and humans. | en |
| dc.description.provenance | Made available in DSpace on 2021-07-11T14:49:20Z (GMT). No. of bitstreams: 1 U0001-1008202012280600.pdf: 2198483 bytes, checksum: a664c5c1094af6041266be30f0dc9e76 (MD5) Previous issue date: 2020 | en |
| dc.description.tableofcontents | Table of content 摘要 i Abstract iii Introduction 1 Material and Method 8 C. elegans strain and maintenance 8 Cell culture 8 RNA interference 9 CRISPR genome editing 9 Supplement assay 10 DHOD-1 inhibition assay 10 UV irradiation-induced cell death assay 10 LC-MS sample preparation 11 Statistical analysis 11 Results 12 Knockdown of dhod-1 causes the bugled tail in the grp-1 mutant 12 Knockdown of umps-1 causes the bulged tail and extra hyp8/9 in the grp-1 mutant 13 Knockdown of genes downstream of umps-1 in the de novo pyrimidine synthesis pathway or degradation pathway did not cause the bulged tail in the grp-1 mutant 14 The normal level of UMP, but not OMP, is required for normal cell death during development 15 Disruption of the salvage pathway is insufficient to induce bulged tail in the grp-1 mutant 16 Uracil and uridine supplementation can recover the bulged tail of the pyr-1; grp-1 mutant 17 Uracil needs to be converted to UMP to rescue the bulged tail of the pyr-1; grp-1 mutant 18 The UMP level is reduced in the pyr-1 mutant 19 The uracil supplementation failed to rescue the bulged tail of the grp-1; ced-3 mutant 19 The DHOD-1 inhibitors cause bulged tail in the grp-1 background 20 Knockdown of mitochondrial UMP transporters does not cause the bulged tail 20 HeLa cell shows decreased percentage of apoptotic cell, after its CAD is knocked down 21 Discussions 23 UMP induced cell death 23 The R12E2.11 RNAi failed to induced the bulged tail in grp-1 mutant 24 The UMP synthesis preference during the embryonic stage 24 The uracil supplementation in NGM or in agar has different efficiency 25 The mitochondrial UMP level is not relevant to PCD 26 The mechanism of UMP caused cell death 26 Figures 27 Figure 1. The scheme of the pyrimidine metabolism. 28 Figure 2. The dhod-1 RNAi caused the bulged tail in the grp-1 mutant 29 Figure 3. The knockdown of ump-1 but not R12E2.11 caused the bulged tail in grp-1 mutant 31 Figure 4. Knockdown of the genes downstream of UMP in de novo pyrimidine synthesis and genes of UMP degradation pathway did not cause the bulged tail in the grp-1 mutant 33 Figure 5. umps-1(ΔOMPDC) caused bulged tail in grp-1 background 34 Figure 6. Knockdown of gene of pyrimidine salvage pathway did not caused the bulged tail in the grp-1 background. 35 Figure 7. Uracil and uridine supplementation rescued the bulged tail of the pyr-1; grp-1 mutant 36 Figure 8. The uracil depletion was not the cause of the ectopic hyp8/9 cell survival 37 Figure 9. The uridine depletion was not the cause of the ectopic hyp8/9 cell survival 38 Figure 10. The UMP level is reduced in the pyr-1 mutant by LC-MS spectrometry 39 Figure 11. The uracil supplementation failed to recue the bulged tail of the grp-1; ced-3 mutant 40 Figure 12. DHOD-1 inhibitor induced the bulged tail in grp-1 mutant 41 Figure 13. Impaired mitochondrial UMP transporter did not cause the bulged tail 43 Figure 14. HeLaCADKD showed less apoptotic cell after exposed to UV irradiation than HeLaScrambled did. 44 References 45 | |
| dc.language.iso | en | |
| dc.subject | 嘧啶生合成 | zh_TW |
| dc.subject | 尿苷單磷酸 | zh_TW |
| dc.subject | 紫外線照射 | zh_TW |
| dc.subject | 代謝物 | zh_TW |
| dc.subject | 計畫性細胞凋亡 | zh_TW |
| dc.subject | UV-irradiation | en |
| dc.subject | metabolite | en |
| dc.subject | pyrimidine de novo synthesis | en |
| dc.subject | programmed cell death | en |
| dc.subject | uridine-5′-monophosphate | en |
| dc.title | 單磷酸尿苷缺乏造成線蟲細胞不正常存活之研究 | zh_TW |
| dc.title | Deficiency of UMP causes inappropriate cell survival in C. elegans | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 108-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 廖秀娟(Hsiu-Chuan Liao),陳昌熙(Chang-Shi Chen) | |
| dc.subject.keyword | 代謝物,嘧啶生合成,計畫性細胞凋亡,尿苷單磷酸,紫外線照射, | zh_TW |
| dc.subject.keyword | metabolite,pyrimidine de novo synthesis,programmed cell death,uridine-5′-monophosphate,UV-irradiation, | en |
| dc.relation.page | 46 | |
| dc.identifier.doi | 10.6342/NTU202002782 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2020-08-31 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 分子與細胞生物學研究所 | zh_TW |
| dc.date.embargo-lift | 2025-08-31 | - |
| 顯示於系所單位： | 分子與細胞生物學研究所 | |
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