2013-08-012024-05-14https://scholars.lib.ntu.edu.tw/handle/123456789/656425摘要：細胞凋亡為一演化上具有高度保留性的細胞死亡機制，對於胚胎發育及維持組織恆定性 非常重要。此機制的失調會導致組織形態上的異常及許多疾病的發生，包括癌症、自體 免疫疾病及神經退化疾病。目前已發現許多調控此機制之分子，例如Bcl-2家族分子就在 調控以粒腺體為主之細胞凋亡過程中扮演了極重要的角色。哺乳動物中，已發現有超過 35個的Bcl-2家族分子，分別調控著細胞以進行死亡（例如：Bax, Bak, Bim等）或抑制死亡 (例如:Bcl-2, A1等)。我們近期發現了一個新的Bcl-2家族分子，將之命名為” Blm-s (Bcl-2 like molecule, small transcript, accession number DQ869383)”，並發現其在哺乳類動物之神 經發育系統中有促進細胞死亡之功能。根據我們初步的結果，發現Blm-s會經由粒腺體釋 放cytochrome c的路徑來調控細胞凋亡，並且在小鼠大腦皮質發育過程中具有促進 postmitotic migratory neuron死亡的功能。我們更進一'步發現Blm-s在經由DNA雙股斷裂的 不可逆刺激下其RNA的表現會被誘發，相較於其他刺激則無此反應。然而，具DNA雙股 斷裂之基因毒性的刺激是如何誘導Blm-s的DNA轉譯表現則尚未被深入研究，因此我們想 在此探討這個誘導的機制（Specific aim-1)。除此之外，Blm-s亦可被tyrosine及serine激酶 磷酸化，且此磷酸化的狀態會影響Blm-s促進細胞凋亡的結果，表示Blm-s可藉由轉譯後修 飾來調控細胞凋亡。然而，我們尚未找到調控Blm-s磷酸化的激酶或去激酶及其被磷酸化 的位置（Specific aim-Il)。許多的Bcl-2家族分子在細胞中的表現位置也影響著它們調控細 胞凋亡的能力，Blm-s亦是如此，我們發現其促進細胞凋亡需要其座落至粒腺體或内質 網。於是我們想找出Blm-s是如何座落至此兩胞器，以及此兩胞器的交互連結如何被Blm-s 影響，並探討Blm-s是否在該胞器上影響其調控細胞内Ca離子濃度進而調控細胞凋亡 (Specific aim-IIl)我們亦計畫產生Mu-s的基因别除鼠來探討Blm-s的生物性功能（Specific aim-IV)。此Mn-s基因剔除鼠若成功產出，可茲用於其他子計畫來探討Blm-s與其他Bcl-2 家族分子或調控細胞凋亡之重要分子如CIP1, ARMS之關係，並運用於病理疾病之研究。<br> Abstract: Apoptosis, an evolutionarily conserved cell-death process, is crucial for normal embryodevelopment and the maintenance of tissue homeostasis. Dysregulation of apoptosis underliesmorphogenetic abnormalities and many diseases including cancer, autoimmune disease andneurodegenerative disorders. Many components of the death machinery have been identified,among which the Bcl-2 superfamily plays a pivotal role as a mitochondria gate-keeper forensuing cell death. In mammals, more than 35 members of Bcl-2 superfamily have beenidentified, and each contributes to the regulation of apoptosis either through pro-apoptotic ability(e.g. Bax, Bak, Bim etc.) or anti-apoptotic machinery (e.g. Bcl-2, A1 etc.). By contributing anovel member to this big family clan, we have recently uncovered a novel molecule, Blm-s(Bcl-2 like molecule, small transcript), which contains the conserved BH domain shared by allBcl-2 families and functions as a pro-apoptotic molecule in the mammalian developing nervoussystem (Accession number DQ869383). Our in vitro and in vivo studies have established thatBlm-s mediates apoptosis via a mitochondrial cytochrome c release pathway and functions inpost-mitotic migratory neurons during murine cortical development. Specifically, Blm-s could betranscriptionally induced in response to irreparable DNA double strand breaks (DSBs), but notto other types of cell stress. However, how genotoxic stress is transduced to the transcriptionalup-regulation of Blm-s is unknown. We thus propose to investigate the signaling machinery thattransduces genotoxic stress generated by DSBs toward the regulation of Blm-s transcription(specific aim-I). Besides, Blm-s could be tyrosine- and serine-phosphorylated, and itsphosphorylation status affects its apoptosis-promoting ability, suggesting that post-translationalmodification of Blm-s is involved in the regulation of cell death. However, the specific amino acidresidues responsible for phosphorylated Blm-s, and the kinases or/and phosphatasesmodulating phosphorylated Blm-s in response to cell stress have not been identified (specificaim-II). Like all Bcl-2 family where proper subcellular localization determines theirapoptosis-mediating capability, Blm-s trafficking to the mitochondria and/or ER is required forapoptosis. Specific questions thus issued such as how Blm-s is targeted to the mitochondriaand ER, and how subcellular localization of Blm-s regulate apoptogenic calcium signaling.These questions will be investigated in our proposed specific aim-III. In specific aim-IV, weaim to generate blm-s-KO mouse to delineate the biological significance of Blm-s in vivo.新穎Bcl-2家族成員-Blm-s粒線體細胞凋亡DNA轉譯及轉譯後修飾調控機制DNA雙股斷裂Wm-s基因剔除鼠Blm-sDSBsphosphorylationtranscription regulationsubcellular localizationmitochondria-centric apoptosisknock-out mouse