請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/33158
標題: | 遺傳分析線蟲dapk-1在計劃性細胞死亡過程中的角色 Genetic Analysis of C. elegans dapk-1 in the Programmed Cell Death Process |
作者: | Wei-Chun Wang 王維君 |
指導教授: | 吳益群(Yi-Chun Wu) |
關鍵字: | 線蟲,細胞死亡, C. elegans,programmed cell death,dapk-1, |
出版年 : | 2006 |
學位: | 碩士 |
摘要: | 計畫性細胞死亡 (細胞凋亡) 是由基因嚴密調控的生化路徑,為多細胞生物發育及維持成體細胞數目恆定所必需。在線蟲 (Caenorhabditis elegans) 中,計劃性細胞死亡受egl-1、ced-9、ced-4、及ced-3等基因調控,而此一調控情形在物種演化過程中具有高度保守性。線蟲DAPK-1為人類死亡蛋白激酶 (DAPK) 之同源蛋白。本實驗室之前對於DAPK-1的研究指出此一蛋白在計劃性細胞死亡過程中扮演一正向調控的角色。本論文中,我們更深入地探討dapk-1之突變株:gk219、ju4、ju469、及ju557。在這些突變株中,胚胎發育過程中由計劃性細胞死亡產生之細胞屍體均顯著減少;且這些突變造成細胞屍體數目下降之程度由高至低依序為:ju557、ju469、gk219、ju4。這些突變株在胚胎時期計劃性細胞死亡之進行對於培養溫度敏感;且ju557為一顯性突變,ju469則為隱性突變。 dapk-1突變除了影響胚胎時期之體細胞死亡,突變株gk219和ju469生殖腺中之細胞屍體數目也有顯著下降之情形。為了定出dapk-1在遺傳調控路徑中的位置,我們在gk219及ju469二突變背景中異位過量表現egl-1於觸感神經元。我們發現gk219抑制了由egl-1所導致之細胞死亡,而ju469對egl-1引起之細胞死亡則有增強之作用。異位過量表現ced-4於觸感神經元內時,gk219與ju469也對ced-4所引起之細胞死亡有相似的影響。於gk219背景中,共同表現dapk-1於觸感神經元中可回復egl-1造成之細胞死亡,顯示dapk-1以細胞自發的方式促進細胞死亡。經由免疫染色我們發現dapk-1廣泛地表現於胚胎細胞中,其於細胞質中呈現絲狀分布,且有集中於靠近細胞膜邊緣之情形。根據實驗結果,我們可以推測DAPK-1 參與胚胎時期體細胞以及成體生殖細胞死亡之進行。且DAPK-1蛋白之不同區域可能依細胞組成不同而扮演不同的角色。 Programmed cell death (apoptosis) is a biological process essential for the development and homeostatic maintenance of multicellular organisms. The main genetic pathway of apoptosis governed by the genes egl-1, ced-9, ced-4, and ced-3 has been characterized in the nematode Caenorhabditis elegans and shown to be highly conserved through evolution. DAPK-1, an ortholog of human DAPK, was previously found to be a positive mediator of somatic programmed cell death in our lab. Here we analyzed dapk-1 mutants (gk219, ju4, ju469, and ju557) in detail. These mutants all exhibited reduced numbers of cell corpses generated by cell death with an allelic order: ju557 (strongest), ju469, gk219, and ju4 (weakest). ju557 was a dominant allele, while ju469 was recessive. Furthermore, the embryonic cell death of these mutants was sensitive to temperature alternation. In addition, dapk-1 mutants (gk219 and ju469) also had reduced numbers of germline cell corpses. To position dapk-1 in the genetic pathway, we expressed egl-1 ectopically in the touch neurons in different dapk-1 (gk219 and ju469) mutant backgrounds. We found that gk219 suppressed but ju469 augmented the cell-killing activity caused by egl-1overexpression. A similar result was obtained when ced-4 was overexpressed in the touch neurons in these mutants. Co-expression of dapk-1 in these cells recovered egl-1 induced cell death in gk219, suggesting that DAPK-1 functioned autonomously to promote cell death. Antibodies against DAPK-1 were generated to examine its expression pattern. DAPK-1 was widely expressed during embryogenesis. The protein exhibited a filamentous pattern in the cytoplasm and was enriched near the membrane periphery. Conclusively, DAPK-1 participates in both embryonic and germline cell death and localizes in the cytoplasm in embryonic cells. It is also indicated that this protein may play different roles depending on the cellular subset. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/33158 |
全文授權: | 有償授權 |
顯示於系所單位: | 分子與細胞生物學研究所 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-95-1.pdf 目前未授權公開取用 | 1.08 MB | Adobe PDF |
系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。