血管動脈粥狀硬化為誘發心血管疾病的主因，好發於血管彎曲和分歧處附近，此處血流型態為振盪型剪應力(oscillatory shear stress, OSS)，易引起內皮細胞產生發炎、增生、氧化等反應，所以被認為參與調控血管內壁病變反應；血管內另一種血流型態為脈衝型剪應力(pulsatile shear stree, PSS)，主要存在於直通型的血管處，能夠啟動血管內皮細胞的內部訊息傳遞，以抑制內皮細胞產生病變，因此，流體剪應力被認為是調控血管內發炎以及動脈硬化斑形成之重要因子。將給予不同流體型態剪應力處理的人類主動脈內皮細胞，進行磷酸蛋白質體學分析，可發現在振盪型剪應力作用下，會促使Yin Yang 1 (YY1)蛋白serine 118位置磷酸化增加；反之，在脈衝型剪應力作用下，磷酸化蛋白的量卻減少，因此我們推測YY1蛋白的磷酸化(p-YY1)可能與血管動脈粥狀硬化形成相關。YY1是一個具多生物功能性的轉錄因子，先前研究指出細胞因應環境需求或刺激，促使YY1活化或者抑制下游基因的表現，進而調控許多複雜的生物功能，包括了胚胎發育、細胞分化、細胞凋亡及腫瘤生成等，近年來研究則顯示YY1在心血管疾病方面，亦扮演重要角色，然而，內皮細胞之YY1在人類血管動脈粥狀硬化中是否扮演的角色及其作用機制尚未被研究。YY1本身能直接調控基因表現，亦能和其他轉錄共同活化因子及轉錄共同抑制因子一起作用來執行功能，因此，本研究目的為篩選出與磷酸化的YY1蛋白(YY1S118E)以及無法磷酸化的YY1蛋白(YY1S118A)之間交互作用具差異之作用分子。利用酵母菌雙雜交技術，以YY1S118E為餌，對人類cDNA 基因庫進行篩選，得知zinc finger with KRAB and SCAN domains 4 (ZKSCAN4)蛋白與YY1S118E有較強的交互作用，並進一步藉由流體與細胞實驗證實在不同流體剪應力作用下，將影響p-YY1和ZKSCAN4的交互作用，得知振盪型剪應力可促進p-YY1與ZKSCAN4蛋白結合並活化下游Hdm2分子，進而抑制p53蛋白之表現與調控細胞增生。綜合言之，我們的結果顯示p-YY1可能參與血管動脈粥狀硬化生成。

Atherosclerosis is one of the major cause of cardiovascular disease, which develops preferentially at arterial branches and curvatures. The fluid shear stress in these regions is oscillatory shear stress (OSS) which has been shown to play important roles in modulating various responses involved in endothelial cell (EC) dysfunction such as inflammation, proliferation, and oxidation. The other kind of fluid shear stress is pulsatile shear stress (PSS) which generally occurs in the straight part of vessels and has been shown to elicit signaling transductions to repress EC dysfunction. Therefore, fluid shear stress is considered as an important factor to modulate inflammation and the formation of atherosclerosis plaques in vascular wall. Through the phosphoproteomic analysis of OSS- and PSS-stimulated human aortic ECs (HAECs), we found that OSS induced the serine 118 phosphorylation of yin yang 1 (YY1) whereas PSS repressed the phosphorylation. Thus, we speculate that the YY1 phosphorylation (p-YY1) may contribute to the development of atherosclerosis. YY1 is known as the ubiquitous transcription factor which possesses multiple biological functions. Previous studies have demonstrated that YY1 exerted its functions by activating or repressing gene expression in response to the developmental cues or stimulations from the surrounding microenvironment. The biological functions YY1 involved include embryogenesis, cell differentiation, cell apoptosis, and tumorigenesis. Recently, it has also been reported that YY1 plays critical roles in the cardiovascular diseases. However, the roles and molecular mechanisms of endothelial YY1 in regulation of atherogenesis are still unstated. YY1 can modulate gene expression through directly acting on promoter regions or interacting with transcriptional co-activators or co-repressors to function. Thus, the aim of this study is to identify molecules which have the differential binding activity with phosphorylated YY1 mimicker (YY1S118E) and YY1 phosphorylation blocker (YY1S118A). By utilizing yeast two-hybrid screening technique which YY1S118E was used as bait and human cDNA library as prey, we identified that zinc finger with KRAB and SCAN domains 4 (ZKSCAN4) has stronger binding activity with YY1S118E. In vitro flow apparatus and cell experiment were further used to study the effect of differential flow pattern on the interaction between p-YY1 and ZKSCAN4. OSS was found to induce the association of p-YY1 and ZKSCAN4, which then activated the expression of downstream gene Hdm2 and in turn, inhibited p53 signal to facilitate the cell proliferation. Altogether, our data demonstrated that p-YY1 may involve in the development of atherosclerosis.

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