2026-11-092026-11-09https://scholars.csmu.edu.tw/handle/123456789/38565Protein arginine methylation has become one of the most intensively studied posttranslational modification (PTM) for cell signaling, protein localization, DNA repair and transcriptional regulation. By now eleven protein arginine methyltransferase (PRMT) genes responsible for the modification have been identified in mammals. Our previous analyses showed that orthologues of mammalian PRMT genes are present in zebrafish, a model organism amenable to genetic manipulations and widely used in developmental studies. In this project we are going to use zebrafish as a model system to elucidate the modification catalyzed by three type I members in the PRMT family in zebrafish early developmental stages. Our major target will be the predominant and ubiquitously expressed PRMT1, its highly conserved paralogue PRMT8 and PRMT6, the only vertebrate PRMT that can automethylate and is intronless. In our ongoing NSC project we have studied the expression pattern of the PRMT genes in different developmental stages of zebrafish, and are dissecting automethylation activity of PRMT6 in human and zebrafish. We are going to use antisense morpholino-oliogonucleotides to specifically inhibit the expression of certain PRMT genes in zebrafish to observe the effects of PRMT knockdowns on early developmental stages. In this new three-year project, we will continue to investigate the physiological roles of PRMT by rescue the morphants. We also propose to set up the proteomic analysis platform of zebrafish to convert to a model system for studying posttranslational modification, specifically, arginine methylation. We will identify the proteins that can be arginine methylated by these PRMT and the distribution, regulation and function of the modifications. Thus in this project, we are going to 1. Continue the PRMT knockdown experiments and perform complement experiments with homologous PRMT or other related genes. Confirm the PRMT physiological functions by rescue experiments of the PRMT morphants. 2. Develop the platform to study zebrafish proteomics and to identify specific methylarginine containing proteins at early embryonic stages 3. Proteomic analyses of the PRMT morphants for the specific methylaccepting substrates of PRMT in zebrafish. 4. Detailed analyses of methylation status of specific methylarginine containing proteins during zebrafish embryonic developments. In this way we will be able to contribute valuable informations of the PRMT genes studied in this project, especially more complete pictures of their functions in a living organism.蛋白質精胺酸甲基化近年來已成為一深入研究的轉譯後修飾，和其相關的細 胞功能包括訊息傳遞、蛋白質胞內定位分佈、DNA 修補以及轉錄調控等。至今已有 十一種人類的蛋白質精胺酸甲基轉移脢 (PRMT) 基因被鑑定出來；我們在胚胎發 育研究的模式生物斑馬魚中找到這些基因的異種同源基因，因此在本計畫中我們將 以斑馬魚為模式生物系統來闡析三種第一型PRMT。我們選擇最主要表現的 PRMT1，其同種同源基因PRMT8 以及可自我甲基化且無intron 的PRMT6 來研究。 我們目前執行中的計畫已研究各種PRMT 於斑馬魚不同發育階段的表達，並將用 morpholino 寡核.酸來抑制個別PRMT 基因表現；我們將觀察knock-down 胚胎早 期變化。在本計畫中我們將研究這些演化上相關的PRMT 基因是否功能上可以部 分互補，並提出以蛋白質體學以及生物資訊的工具將斑馬魚轉變為一個研究轉譯後 修飾主要精胺酸甲基化的模式系統。在本計畫中我們會 1. 持續用morpholino 寡核.酸來抑制個別PRMT 基因表現；我們將觀察 knock-down 胚胎早期變化，以打入相關基因的mRNA 救援實驗來確定所 觀察外表型變化及所推測的生理功能途徑。 2. 建立斑馬魚蛋白質體研究平台鑑定胚胎發育早期含精胺酸甲基之蛋白 3. 以蛋白質體研究平台鑑定斑馬魚PRMT morphants 含精胺酸甲基之蛋白。 4. 詳盡分析某些特定的精胺酸甲基接受蛋白及其甲基化於胚胎發育早期之重 要性。 預期本計畫之進行後，我們將可對這幾種甲基轉移脢提供寶貴資訊，特別是 在一生物活體中其功能的完整呈現。Project