臺灣博碩士論文加值系統

果蠅的外在神經器官 (external sensory organ) 由五個不同的細胞所組成：毛細胞 (hair cell)、毛窩細胞 (socket cell)、神經細胞 (neuron)、神經鞘細胞 (sheath cell) 和神經膠細胞 (glia cell)。這五個細胞由同一個感覺器官先驅細胞 (sensory organ precursor) 經過四次的不均等細胞分裂 (asymmetric cell division) 所產生。此過程需要細胞命運決定物質Numb，在細胞分裂時的不均等分佈及Notch訊號在兩個子細胞的不均等激發。Nak (Numb-associated kinase) 是用Numb做釣餌由酵母雙雜交篩選 (yeast two-hybrid screen) 所分離出來。前人研究顯示Nak為一200kD的蛋白並具有Ser/Thr激酉每區，在果蠅活體內大量表現時可以拮抗Numb的功能。
為了進一步瞭解Nak在果蠅活體內的功能，我製做了幾個可以增加或減少與Numb之親和力，或激酉每區加入突變的nak轉殖基因果蠅品系。由測試這些轉殖基因影響果蠅感覺器官發育的結果顯示，激酉每活性或與Numb的親和力對於Nak的功能是重要的。此外大量表現Nak並不干擾Numb在神經前驅細胞 (neuroblast) 的不均等分佈，所以Nak對於Numb功能上的拮抗應該作用於細胞命運決定的時期。
另一方面，我分析了三個可能為nak突變的基因性狀（nak1, nak2 and nak3）。一般而言，這些突變的同源合子會死於幼蟲初期，而且有細胞增殖的問題。透過對鑲嵌突變動物和表達Nak於特定組織的性狀分析，Nak可能在眼睛分化、翅緣與翅脈的形成、組織形變等方面參與作用。這些結果顯示Nak可能是Notch訊號途徑的正向調控因子而且可能參與細胞骨骼的調控。

The Drosophila external sensory organ is composed of five cells, a socket cell, a hair cell, a sheath cell, a neuron and a glia cell, which are derived from a single sensory organ precursor (SOP) cell. Generation of distinctive cell fates in the sensory organ lineage is achieved by four rounds of asymmetric cell divisions that require unequal distribution of the cell fate determinant Numb in the mitotic cell and differential activation of Notch signaling in two sibling cells. In a previous study, Nak (Numb-associated kinase) was isolated in a yeast two-hybrid screen using Numb as a bait. nak encodes a 200kD protein with a Ser/Thr kinase domain, and by misexpression Nak is able to antagonize the function of Numb in vivo.
To understand the nak's function in vivo, I constructed several nak transgenes which either increase or decrease the affinity with Numb, or in combination with mutations in the kinase catalytic domain. These transgenes were germ-line transformed to generate transgenic flies and their effects in sensory organ development were assayed. My results suggested that both kinase domain and affinity to Numb are important for nak's function in vivo. In addition, nak overexpression in neuroblasts fails to disturb the normal localization of Numb, indicating that nak might antagonize the function of Numb in the step of cell fate determination.
In another approach, I analyzed mutant phenotypes from three nak candidate alleles (nak1, nak2 and nak3) isolated in the lab. In general, nak homozygous animals were larval lethal and were defective in cell proliferation. Analysis of mutant mosaic clones and overexpression of Nak in specific tissues were performed to investigate the endogenous role of Nak. In summary, Nak may play roles in eye differentiation, wing margin and vein formation, and tissue morphogenesis. Our results suggest that Nak could be a positive regulator of the Notch pathway and might participate in cytoskeleton regulation.

中文摘要 1
Abstract 2
Introduction 4
Materials and Methods 7
Results 11
Discussion 17
References 22
Figures and Table 26
Appendix 38

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