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研究生:楊雯婷
研究生(外文):Yang, Wen-Ting
論文名稱:Cadherins mediate cell sorting
論文名稱(外文):Cadherins調控細胞分離現象之研究
指導教授:徐瑞洲
指導教授(外文):Hsu, Jui-Chou
口試委員:陳令儀范聖興徐瑞洲
口試日期:2011-7-26
學位類別:碩士
校院名稱:國立清華大學
系所名稱:分子醫學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2011
畢業學年度:99
語文別:英文
論文頁數:46
中文關鍵詞:鈣黏著分子細胞分離
外文關鍵詞:cadherincell sorting
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Cell sorting is a morphogenetic processthat cell populations with different identities may segregate from each other. Being a major cellular adhesion molecule on adherens junction, Drosophila epithelial cadherin (DE-cadherin), the homologue of E-cadherin in Drosophila, can mediate cell-cell adhesion through the extracellular domain that interacts with other cadherins and through the intracellular domain that forms the cadherin-catenin complex. Alterations in DE-cadherin expression levels may lead to cell sorting by changing the adhesivity. On the purpose to investigate how cadherins mediate cell sorting, I overexpressed cadherins, the truncated forms and the chimera cadherins under the control of GAL4-UAS system in Drosophila wing discs and subsequently observed the phenotypes in a macro view. The results in this study revealed that a full-length of cadherin was required for the mediation of cell sorting behavior. Both the extracellular domain and the intracellular domain of a cadherin were required, however, not sufficient while acted alone, for cell sorting. Signals from cell-cell contact had to transmit to the cytoskeleton through interactions of the cadherin-catenin complex, cooperatively, to mediate the cell sorting behavior of cell populations.
細胞分離(cell sorting)是一種形態生成的過程當中常見的現象:性質不相同的細胞會傾向與彼此分離;而使得性質接近或相同的細胞聚集在一起。果蠅的上皮鈣黏著分子(Drosophila epithelial cadherin, DE-cadherin)是細胞黏著分子(cellular adhesion molecule)的一種,可以透過其細胞外的結構與其他上皮鈣黏著分子進行順式或反式的結合,並同時藉由其細胞內結構與連環蛋白(catenin)形成複合物,以調控細胞與細胞之間的連結。改變上皮鈣黏著分子的表現量會影響細胞間的黏著作用,因而導致了細胞分離現象的產生。為了瞭解上皮鈣黏著分子是如何調控細胞分離的現象,在這個研究當中我利用了GAL4-UAS system將全長、截短以及嵌合等形態的鈣黏著分子大量表現在果蠅的翅碟(wing disc)上,並且以巨觀的角度探討鈣黏著分子對於細胞分離現象的調控。實驗結果顯示,只有全長的鈣黏著分子得以調控細胞分離的現象。單就細胞外結構或是細胞內結構的表現量改變是不足以使細胞群分離開來的,兩者必須共同作用,使得細胞與細胞接觸時的訊息藉由鈣黏著分子與連環蛋白所形成的複合物進一步地傳遞至細胞骨架(cytoskeleton),以調控細胞分離的行為與現象。
Abstract
1 Introduction….……………………………………….........................………- 1 -
2 Methods and Materials……………………………………………………....- 3 -
2.1 Drosophila stocks………………………………………………………..- 3 -
2.2 Molecular cloning...……………………………………………………..- 3 -
2.2.1 pUAST-DEcadΔcyto-DNcadintra-HA (DEDN-cadherin) …………………………………………………..- 3 -
2.3 Histochemistry…………………………………………………………..- 4 -
2.4 TUNEL assay……………………………………………………………- 5 -
2.5 Cuticle specimen preparation…………………………..………………..- 5 -
3 Results………………………………………………………………………....- 7 -
3.1 Alteration in DE-cadherin expression level led to deformation of wing imaginal discs……………………………………………………..……..- 8 -
3.1.1 Overexpression of DE-cadherin………………………...…………..- 8 -
3.1.2 Down-regulation of DE-cadherin…………………………………...- 9 -
3.2 Ectopic expression of DE-cadherin ECD and DE-cadherin ICD had no effects on morphology of wing imaginal discs………………………...- 10 -
3.3 Ectopic expression of DN-cadherin led to deformation of wing imaginal discs……………………………………………………………..……....- 11-
3.4 Ectopic expression of DEcadΔcyto-DNcadintra-HA led to deformation of wing imaginal discs………………………………………………………......- 12 -
3.5 Ectopic expression of DE-cadherin Δβ had no effects on morphology of wing imaginal discs………………………………………………….....- 14 -
3.6 Ectopic expression of DE-cad Δcyto-α-catenin had no effects on morphology of wing imaginal discs………………………………………………....- 15 -
3.7 Summary………………………………..……………………………....- 16-
4 Discussion………………………………………………………………........- 17 -
4.1 ECDs and ICDs of cadherins cooperatively mediate cell sorting……...- 17 -
4.2 Overexpression of cadherin ICDs leads to the notch phenotype in wing blades independent of cell sorting……………………………………...- 19 -
4.3 Cells ectopically expressing cadherins are restricted in a small area…..- 21 -
5 Figures…………………………………………………………….……........- 25 -
5.1 Figure 1. Wild type, truncated form and chimeras of cadherins used in this study……………………………………………………………............- 25 -
5.2 Figure 2. The expression patterns of wingless (wg) and different Gal4 lines used in this study……………………………………………………….- 26 -
5.3 Figure 3. Overexpression of DE-cadherin leads to deformation of wing discs…………………………………………………………….............- 27 -
5.4 Figure 4. Overexpression of DE-cadherin has no significant correlation with cell death……………………………………………….................- 28 -
5.5 Figure 5. RNA interference of DE-cadherin leads to deformation of wing discs……………………………………………….................................- 29 -
5.6 Figure 6. Ectopic expression of DE-cadherin ECD and DE-cadherin ICD does not affect the morphology of wing discs…………........................- 30 -
5.7 Figure 7. Phenotypes in notum and wing blade resulted from ectopic expression of DE-cadherin ICD, DN-cadherin and DEcadΔcyto-DNcadintra-HA…………………….......................................- 32 -
5.8 Figure 8. Ectopic expression of DN-cadherin leads to deformation of wing discs…………………….........................................................................- 34 -
5.9 Figure 9. Ectopic expression of DEcadΔcyto-DNcadintra-HA leads to deformation of wing discs…………………….......................................- 36 -
5.10 Figure 10. DEDN-cad-expressing cells are expelled from the region of wing pouches……………………...........................................................- 38 -
5.11 Figure 11. Ectopic expression of DE-cadherin Δβ does not affect the morphology of wing discs.......................................................................- 39 -
5.12 Figure 12. Ectopic expression of DE-cadherin-α-catenin does not affect the morphology of wing discs.......................................................................- 40 -
5.13 Figure 13. Ectopic expression of DE-cadherin-α-catenin does not affect the distribution of F-actin..............................................................................- 41 -
6 References........................................................................................................- 42 -


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