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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 吳益群 | |
dc.contributor.author | Po-Hsuan Li | en |
dc.contributor.author | 李柏萱 | zh_TW |
dc.date.accessioned | 2021-06-08T00:44:28Z | - |
dc.date.copyright | 2015-08-10 | |
dc.date.issued | 2015 | |
dc.date.submitted | 2015-08-06 | |
dc.identifier.citation | Armenti, S.T., and Nance, J. (2012). Adherens junctions in C. elegans embryonic morphogenesis. Subcell Biochem 60, 279-299.
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/17839 | - |
dc.description.abstract | 表皮系統(epidermis systems)位於生物體的最外層,表皮細胞之間存在細胞連結(cell-cell junctions)分子,將表皮細胞緊密的排列再一起,可作為生物體與外界環境的保護屏障,以避免受到外來物質的入侵。線蟲的表皮接縫細胞(seam cells) 直線排列在身體的左右兩側,每側各有16顆接縫細胞,並且鑲嵌於圓柱形的hyp7合胞體(syncytium)細胞間,由黏著連接分子(adhesion molecules)將接縫細胞與hyp7皮下細胞連接在一起。在第四幼蟲時期,線蟲的16個接縫細胞會融合成一個多核的合胞體(syncytium),我們利用AJM-1::GFP或HMR::GFP的螢光融合蛋白來標記連接分子,可以觀察到在成蟲的細胞邊界會形成平行排列的兩條線,但在blmp-1基因缺失的成蟲中,會觀察到類似於泡泡狀的異常細胞型態。利用分布在表皮底層的分子蛋白,發現泡泡狀的異常現象只存在於表皮細胞的上層區域,而細胞底區域是正常的。經不同時間的螢光顯微照相追蹤發現 ,blmp-1突變株在幼蟲蛻皮剛進入成蟲階段,接縫細胞邊界變得不規則,兩條邊界的距離縮短,並且會產生交接點。因此blmp-1基因在維持成蟲表皮細胞型態扮演重要的角色,在hyp7或接縫細胞降低blmp-1活性皆可造成泡泡狀的接縫細胞形態,顯示blmp-1在hyp7與接縫細胞中都不可或缺的。blmp-1是一個轉錄抑制因子(transcription repressor),相似於哺乳類中的PRDI-BFI/blimp-1(B lymphocyte-induced maturation protein 1),因此我們篩選出含有BLMP-1 binding domain且在blmp-1缺失的線蟲中其表現量較高的基因,利用RNAi knockdown的方式,試圖找出降低這些候選基因的表現可以下降成蟲異常表皮細胞型的比例的產生,並發現在blmp-1突變的線蟲中抑制bus-8的表現,可以大幅度的降低異常表皮細胞的型態。BUS-8是一個參與醣化作用的蛋白,且表現在接縫細胞內。另外,發現其他醣化作用的蛋白bus-2,bus-4和bus-12也可以下降部分異常表皮細胞型態。因此我們認為blmp-1會透過bus-8或其他醣化作用蛋白,來維持成蟲表皮細胞的型態。 | zh_TW |
dc.description.abstract | Epithelial cells are linked by cell-cell junctions that hold the tissue together, and function as a protective barrier. In the nematode Caenorhabditis elegans, epidermal seam cells are arranged in longitudinal rows on the left and right sides of the body and are embedded in the cylindrical hyp7 syncytium. Seam cells and hyp7 are connected by adhesion junctions along their apical borders. These cell junctions are essential for cell polarity, adhesion and innate immunity. In the 4th larval (L4) stage, seam cells become terminally differentiated and are fused to make one syncytium of 16 cells on each side. Using AJM-1::GFP and HMR-1::GFP fusion proteins that mark the apical adhesion junction, I observed two almost parallel lines that run along the seam syncytium boundary on each side of the body in the wild-type adult animals. However, in mutants defective in blmp-1, which encodes a zinc finger transcription factor similar to mammalian transcriptional repressor BLIMP-1(B lymphocyte-induced maturation protein 1), the apical epithelial junctions showed a bubble-like arrangement in seam cells in adult. By performing the time-course analysis of the AJM-1::GFP pattern in hypodermis, I found that, in the blmp-1 mutant, seam cell fusion and the adhesion junction arrangement is normal in L4, but the border between seam cell and hyp7 became irregular and the opposite sides of the border started became attached at multiple points during the L4/adult molt. The epidermal basolateral region marker LET-413::GFP showed no detectable abnormality in the basal region of the seam syncytium. In addition, inactivation of blmp-1 by RNA interference in either seam or hyp7 resulted in bubble-like apical epithelial junctions, showing that blmp-1 is essential in both seam and hyp7 to maintain the normal apical surface of the seam syncitium. Since BLMP-1 can function as a transcriptional repressor, this seam cell apical surface defect may be caused by abnormally high expression of some target genes. Using candidate genes approach and a function test, I found that bus-8 RNAi significantly reduced the percentage of blmp-1 mutants with the abnormal apical seam cell shape. BUS-8 is predicted as a mannosyltransferases involved in protein glycosylation, such as bus-2, bus-4, bus-12, partially rescued the apical seam cell shape defect of the blmp-1 mutant. On the basis of these data, I proposed that loss of blmp-1 caused the abnormality of apical seam cell shape due to abnormal protein glycosylation by BUS-8, in particular, and BUS-2, BUS-4 and BUS-12, in part, in seam and/ or hyp7 cells and that proper glycosylation of extracellular proteins or membrane proteins is important for the maintenance of the apical epithelial cell shape. | en |
dc.description.provenance | Made available in DSpace on 2021-06-08T00:44:28Z (GMT). No. of bitstreams: 1 ntu-104-R02b43002-1.pdf: 2506219 bytes, checksum: d5c78f8cb507298074b8cc3f61817296 (MD5) Previous issue date: 2015 | en |
dc.description.tableofcontents | 口委審定書……………………………………………………………………………...i
致謝 ……………………………………………………………………………………ii 中文摘要 ………………………………………………………………………………iii Abstract …………………………………………………………………………………v Introduction …………………………………………………………………………….9 Materials and Methods ………………………………………………………………...16 1) C. elegans strains ……………………………………………………………16 2) Plasmid construction ………………………………………………………..17 3) Microscopy …………………………………………………………………18 4) Microinjection ………………………………………………………………19 5) RNAi experiment ……………………………………………………………19 Results …………………………………………………………………………………21 1) blmp-1 mutants displayed an abnormal AJM-1::GFP at the early adult stage 21 2) blmp-1 mutants also displayed a bubble-like HMR-1::GFP pattern ………..22 3) The abnormal seam cell boundary of blmp-1 mutant is in the apical but not basal region. ………………………………………………………………………22 4) The bubble-like structure formation happens between L4/adult molt and young adult stages, and this process is dynamic. ………………………………...23 5) blmp-1 is expressed in both hyp7 and seam cells at late L4 and adult stages. …………………………………………………………………………….24 6) Knockdown of blmp-1 in seam or hyp7 leads to abnormality in apical epithelial shape in adult animals. …………………………………………………24 7) Down-regulation of bus-8 expression significantly suppress the abnormal bubble-like apical epithelial shape in blmp-1 adult mutant ………………………25 8) blmp-1(s71); bus-8(e2885) mutant displays the normal arrangement of apical epithelial shape in seam syncytium. ……………………………………………...26 9) Other glycosylation genes, which bus-2, bus-4, and bus-12, partially rescue the abnormality of apical epithelial shape in blmp-1(RNAi) animals. …………...27 10) blmp-1 mutant displays the short width of seam syncytium at the late L4 stage. ……………………………………………………………………………...28 References ……………………………………………………………………………..33 Figures …………………………………………………………………………………38 Figure 1. Seam cells are arranged as longitudinal rows of 16 cells on the left and right sides of the body at the L4 and adult stages, and they are embedded in the cylindrical hyp 7 syncytium. ……………………………………………………..36 Figure 2. Seam cell lineages and development. …………………………………..37 Figure 3. The C. elegans apical junction and analogous junctions in Drosophila and mammals ……………………………………………………………………..38 Figure 4. Gene structure of blmp-1. ……………………………………………...39 Figure 5. Seam cell development during L4 and adult stages. …………………..40 Figure 6. blmp-1 mutant animals show a bubble-like pattern at the adult stage …41 Figure 7. Models of seam syncytium shape in wild-type and blmp-1 mutant. …..42 Figure 8. The abnormal seam syncytium boundary of blmp-1 mutant may occur at apical but not basal region. ………………………………………………………43 Figure 9. Seam cell boundary becomes irregular, and the process of bubble-like structure formation is dynamic in blmp-1 mutant. ……………………………….44 Figure 10. Two AJM-1::GFP lines between seam and hyp7 do not touch each other in a wild type animal. …………………………………………………………….45 Figure 11. blmp-1 is expressed in both hyp7 and seam cells at late L4 and young adult stages……………………………………………………………………… 46 Figure 12. Quantification of worms that showed the abnormalbility of apical epithelial shape by cell-specific blmp-1 RNAi ………………………………….47 Figure 13. Quantification of worms that showed the abnormalbility of apical epithelial shape in seam cell by feeding different genes RNAi in blmp-1 (A) and wild type (B) animals. ……………………………………………………………49 Figure 14. blmp-1(s71); bus-8(e2885) double mutant animals display no defect of apical epithelial shape in seam syncytium ……………………………………….50 Figure 15. bus-8 is expressed in seam cells ………………………………………51 Figure 16. Model of BLMP-1 regulating apical epithelial shape through glycosyltransferase BUS-8 ……………………………………………………….52 Tables …………………………………………………………………………………..53 Table 1. blmp-1 mutations caused defective seam syncyitum phenotype. ……….53 Table 2. The candidate genes of blmp-1 target genes …………………………….53 Table 3. bus-2, bus-4, bus-12 also partially rescue the abnormal phenotype in blmp-1(RNAi) animals. …………………………………………………………..54 Table 4. The width of seam cells …………………………………………………54 Supplementary data ……………………………………………………………………55 S1. Actin arrangement in blmp-1(RNAi)animals. ………………………………..55 | |
dc.language.iso | en | |
dc.title | BLMP-1經由醣基轉移酵素BUS-8調控線蟲上層表皮細胞的形狀 | zh_TW |
dc.title | C. elegans BLMP-1 regulates apical epithelial shape through glycosyltransferase BUS-8 | en |
dc.type | Thesis | |
dc.date.schoolyear | 103-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 陳昌熙,潘俊良,徐瑞洲 | |
dc.subject.keyword | 細胞形狀,表皮細胞,blmp-1,bus-8,接縫細胞,醣化作用, | zh_TW |
dc.subject.keyword | cell shape,epidermis,blmp-1,bus-8,seam sycytium,glycosylation, | en |
dc.relation.page | 55 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2015-08-07 | |
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 |
顯示於系所單位: | 分子與細胞生物學研究所 |
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