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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 蔡欣祐 | zh_TW |
| dc.contributor.advisor | Hsin-Yeu Tsai | en |
| dc.contributor.author | 吳念祖 | zh_TW |
| dc.contributor.author | Nien-Tsu Wu | en |
| dc.date.accessioned | 2024-08-19T16:23:50Z | - |
| dc.date.available | 2024-08-20 | - |
| dc.date.copyright | 2024-08-19 | - |
| dc.date.issued | 2024 | - |
| dc.date.submitted | 2024-08-10 | - |
| dc.identifier.citation | 1. Song, J.-J., et al., Crystal structure of Argonaute and its implications for RISC slicer activity. science, 2004. 305(5689): p. 1434-1437.
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Pugacheva, E.M., et al., BORIS/CTCFL epigenetically reprograms clustered CTCF binding sites into alternative transcriptional start sites. Genome Biology, 2024. 25(1): p. 1-40. 32. Heger, P., B. Marin, and E. Schierenberg, Loss of the insulator protein CTCF during nematode evolution. BMC molecular biology, 2009. 10: p. 1-14. 33. del Castillo-Olivares, A., M. Kulkarni, and H.E. Smith, Regulation of sperm gene expression by the GATA factor ELT-1. Developmental biology, 2009. 333(2): p. 397-408. | - |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/94784 | - |
| dc.description.abstract | 小RNA在各種物種的精子功能中起著至關重要的作用。在秀麗隱桿線蟲（Caenorhabditis elegans）中，缺乏精子相關的小RNA「alg-3/4 26G小RNA」會導致線蟲在25攝氏度時不育，然而，大多數先前的研究集中在缺乏alg-3/4 26G小RNA的病理影響上，其上游調控機制仍不清楚。因此我們的研究深入探討了26G sRNA的生物合成機制，針對類zc3h12a核糖核酸酶NYN-3如何辨識mRNA模板並進行切割進行研究。欲探討26G小RNA生成機制，我們分析了ALG-3結合的mRNA中的序列元素和轉錄起始位點（TSS），提出NYN-3的識別並不依賴特定序列，而是依靠結合轉錄起始位點的蛋白進行辨識。
根據文獻探討，我們發現線蟲中真核翻譯起始因子IFE-1作為結合轉錄起始位點的蛋白，非常有可能協助NYN-3辨認mRNA模板。IFE-1是人類真核翻譯起始因子EIF4E的直系同源物，主要表達於雄性生殖細胞系統中，並表現於雄性生殖腺的近端區域，這與NYN-3和ALG-3的基因表現相一致。我們的研究結果發現IFE-1在雄性生殖腺的近端區域與NYN-3短暫相互作用，推斷IFE-1能幫助NYN-3識別mRNA 進而產生26G sRNA合成所需的RNA模板。 | zh_TW |
| dc.description.abstract | Small RNAs play a crucial role in sperm function across various species. In Caenorhabditis elegans, the absence of sperm-related small RNAs, alg-3/4 26G small RNAs, results in infertility at 25 degrees Celsius. Most prior studies focused on the pathological effects of missing alg-3/4 26G small RNAs, the upstream regulatory mechanisms have remained unclear. Our research delves into the mechanisms of 26G sRNA biogenesis, particularly the targeting rules of the zc3h12a-like ribonuclease NYN-3, which is essential for the cleavage of mRNA templates required for 26G sRNA synthesis. We investigate the sequence elements and transcription start sites (TSSs) in ALG-3-bound mRNAs, proposing that NYN-3 recognition is independent of specific sequences, but rather involves interactions with TSS-binding proteins. We further identify the eukaryotic translation initiation factor IFE-1 as a potential TSS-binding protein that may assist NYN-3 in 26G sRNA biogenesis. IFE-1, an ortholog of human EIF4E, is predominantly expressed in the male germline and localizes to the proximal region of the male gonad, aligning with the expression patterns of NYN-3 and ALG-3. Our findings demonstrate that IFE-1 transiently interacts with NYN-3 at the proximal region of the male gonad, suggesting a collaborative role in recognizing target mRNAs for 26G sRNA production. | en |
| dc.description.provenance | Submitted by admin ntu (admin@lib.ntu.edu.tw) on 2024-08-19T16:23:50Z No. of bitstreams: 0 | en |
| dc.description.provenance | Made available in DSpace on 2024-08-19T16:23:50Z (GMT). No. of bitstreams: 0 | en |
| dc.description.tableofcontents | Master Thesis Acceptance Certification …………………………………………....i
誌謝………………..........................………………………………………………...ii 摘要…………………………………………….…………………………………..iii Abstract ……………………..………………………….……………………………..iv Table of Contents……………………………..…….……………………………...v Chapter 1. Introduction………………………………………...………………………. 1 1.1 The role of small RNAs in germline development……………………...……... 1 1.2 The regulation of piRNAs during spermatogenesis ……………………2 1.3 The alg-3/4 26G small RNAs specifically regulate spermiogenesis……………2 1.4 The biogenesis of alg-3/4 26G small RNAs ……………………………4 1.5 The potential mechanism NYN-3 targeting alg-3/4 26G small RNAs …………5 1.6 Eukaryotic translation initiation factor in C. elegans……………………....………5 1.7 The potential roles of IFE-1 in 26G small RNA regulation ………………..……6 Chapter 2. Materials and Methods …………………………………………….. 7 2.1 Strains ……………………………………………………………….……………7 2.2 Small RNA library preparation …………………………………………………7 2.3 Verification of gfp small RNA in reporter strain ……………..…………………9 2.4 Detection of alternative transcripts in males and hermaphrodites………………10 2.5 Brood size test …………………………….………………………………...…10 2.6 Worm lysate preparation ………………...………………………………………11 2.7 Immunoprecipitation …………….………...……………………………………..11 2.8 Western blot ……………….…….………...……………………………………..12 2.9 Immunofluorescence staining ………….……….……………………………..13 2.10 Proximity ligation assay ………….……….…………………………………..14 Chapter 3. Results …………………………………………………………………16 3.1 NYN-3 recognition is sequence-independent and doesn’t solely rely on the stem loop structure …………………………………….……………….…………16 3.2 The biogenesis of ALG-3/4 26G sRNA likely depends on transcription start sites ……………………………………………………………..…….…………18 3.3 IFE-1, a germline-specific homolog of EIF4E, potentially aids NYN-3 in recognizing target mRNAs ……….……..………………………….…………19 3.4 IFE-1 transiently interacts with NYN-3 at the proximal region in male gonad……….………………………………………………….…….…………20 Chapter 4. Results ……………………………………………………………………… 23 4.1 Transcription factors and 26G sRNAs both regulate the expression of sperm- specific genes during spermatogenesis ……………………….…….…………23 4.2 The targeting rules of NYN-3 recognizing alg-3/4 targeted mRNA remains to be discussed ………………………………………………….…….…….…………24 4.3 IFE-1 potentially guides NYN-3 recognition of alg-3/4 targeted mRNA for the biogenesis of 26G small RNA …………………………….…….…….…………25 Chapter 5. Figures and Tables ……………………………..…………………………… 27 Figure 1.The gfp small RNAs exist in alg-3/4 small RNA reporter strain…….…….. 27 Figure 2. The discovery of alternative transcripts for alg-3/4 targeted mRNAs.……. 31 Figure 3. Characterization of ife-1 wild type and ife-1 (tm4249) null mutant……... 34 Figure 4. Transient interaction between NYN-3 and IFE-1………………………... 36 Figure 5. Proposed model of the role of IFE-1 in 26G sRNAs biogenesis…………... 41 Tables………………………………………………………………………………... 42 Table-I Primers and adaptors used for small RNA library preparation …………... 42 Table-II Primers for target small RNA detection …………………………………... 42 Table-III Primers for alternative transcripts detection ……………………………... 43 Reference………………………………………………………………………………... 44 | - |
| dc.language.iso | en | - |
| dc.title | 於秀麗隱桿線蟲中探討調控精子之小RNA的生成機制 | zh_TW |
| dc.title | Decoding the Mechanisms Governing Small RNA Biogenesis Essential for Sperm Function in Caenorhabditis elegans | en |
| dc.type | Thesis | - |
| dc.date.schoolyear | 112-2 | - |
| dc.description.degree | 碩士 | - |
| dc.contributor.oralexamcommittee | 林劭品; 詹世鵬 | zh_TW |
| dc.contributor.oralexamcommittee | Shau-Ping Lin;Shih Peng Chan | en |
| dc.subject.keyword | 秀麗隱桿線蟲,精子生成,轉譯起始因子,生殖細胞發育,26G小RNA,ALG-3/4, | zh_TW |
| dc.subject.keyword | C. elegans,Spermatogenesis,Translation initiation factors,germline development,26G small RNAs,ALG-3/4, | en |
| dc.relation.page | 47 | - |
| dc.identifier.doi | 10.6342/NTU202403521 | - |
| dc.rights.note | 同意授權(全球公開) | - |
| dc.date.accepted | 2024-08-12 | - |
| dc.contributor.author-college | 醫學院 | - |
| dc.contributor.author-dept | 分子醫學研究所 | - |
| 顯示於系所單位： | 分子醫學研究所 | |
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