2019-01-012024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/694843摘要：精漿中的精漿蛋白質具有促進受孕率與提高精子活力的功用。本研究團隊經蛋白質譜分析發現，精漿中一個高含量的蛋白質，稱為硫氫氧化&#37238;QSOX（quiescin Q6/sulfhydryl oxidases），具有跨物種凝集不正常精子之特性（例如：已知齧齒類QSOX 蛋白可凝集人與豬之精子），進一步分析更發現QSOX的兩種亞型（isoform）於生殖道的分佈不同，並且，在附睪不同區段中，不同亞型與精子結合的位置也不同，因此，我們推測精漿蛋白質QSOX對於精子之各項功能應有重要影響，並可能具有調節精子功能與穩定精子細胞膜蛋白型態之特性。本計畫預期利用CRISPR/Cas9基因編輯技術與Cre-LoxP 系統建置全基因剔除與器官專一性基因轉殖小鼠，針對QSOX基因與蛋白質在生殖道之功能與其調控機制進行深入的分系與探討。此外，利用建立的QSOX全基因剔除細胞株（QSOX1-/-, QSOX2-/-）及螢光QSOX細胞株 (eGFP-QSOX1, eRGP-QSOX2)，以特殊二維（2D）極性細胞培養模式探討QSOX對精子各項功能之影響及其分子調控機制。有鑑於目前研究QSOX的文獻缺乏，此計畫預期深入了解QSOX基因與蛋白質於生殖道調控精、卵熟程過程的分子調控機制，並可進一步運用此計畫建立的研究系統與方式，廣泛應用在研究其他器官之上皮細胞蛋白質分泌的分子調控機制（例如腎臟上皮細胞調控管腔酸鹼度、輸卵管上皮細胞調控卵子附著與精、卵結合調控）。<br> Abstract: In Taiwan, the total amount of biotechnology spent on the treatment of infertility exceeds NT$20 billion per year; however, in the absence of a comprehensive understanding and evaluation system, causes for infertility problem cannot be fully explained and cure, and as a consequence, techniques facilitate fertilization success cannot be effectively improved. Recent studies demonstrated that proteins within seminal plasma have various functions to promote pregnancy success. We previously identified, through the proteomic analysis, a high level sulfhydroxylase QSOX (quiescin Q6/sulfhydryl oxidases) protein present in the semen, and exhibits a inter-species agglutination of abnormal sperm (eg: rodent QSOX1 can recognize and agglutinates human or pig sperm), further analysis of the QSOX found the existence of two protein isoforms with complementary tissue distribution in the male reproductive tract. Moreover, these isoforms associate at specific, but distinct sperm membrnae surface of maturating sperm cells. Therefore, we hypothesize that the QSOX proteins may play important roles in the regulation of sperm maturation process and its physiological functions. This project will use the CRISPR/Cas9 gene editing technology and the Cre-LoxP system to construct whole-gene knockout and organ-specific transgenic mice, and conduct in-depth systematic research on the function of QSOX genes and proteins in the male reproductive tract. In addition, the use of the established QSOX KO cell line (QSOX1-/-, QSOX2-/-) and fluorescent-tagged QSOX cell line (eGFP-QSOX1, eRFP-QSOX2) for in vitro 2D polarized cell culture system, we can investigate the molecular basis of QSOX effect on sperm function and the underlysing regulatory mechanism. Due to current lack of literature information on QSOX, this project is expected to gain insight into the role of QSOX genes and proteins in the process of sperm maturation, and to explore the feasibility of QSOX gene editing or the stimulation and inhibition of QSOX protein secretion to directly or indirectly affect sperm quality that lead to better fertilization success. The experimental system used in this project will allow further applications on the study of epithelium-cell/gamete interactions in a broad range of organs, for example to study acidification of renal lumen, the molecular regulation of oocyte maturation and sperm-oocyte interaction from the oviduct epithelium.精子精漿蛋白不孕spermatozoaseminal vesicle proteinsinfertilityreproductive tract&#1048779&#1048616精子精漿蛋白不孕spermatozoaseminal vesicle proteinsinfertility