酵母菌第一腺嘌呤核苷二磷酸核醣化因子相似蛋白和高爾基體蛋白Imh1p於高爾基氏體成熟面網上的功能性探討

Functional Characterization of ARF-like protein Arl1p and Golgin protein Imh1p at the trans-Golgi network

陳玟卉(Wen-Hui Chen)

指導教授 ： 李芳仁

國立臺灣大學/醫學院/分子醫學研究所/碩士(2015年)

https://doi.org/10.6342/NTU.2015.10528

摘要

第一腺嘌呤核苷二磷酸核醣化因子相似蛋白 (ARL1) 是腺嘌呤核苷二磷酸核醣化因子 (ADP-ribosylation factors , ARF) 家族的一員，其主要參與在高爾基氏體囊泡運輸的過程。近年來在酵母菌為模式生物研究中，我們實驗室已知道Arl1p的鳥糞嘌呤核苷三磷酸 (GTP) 交換蛋白 (GEF) Syt1p 可以促進 Arl1p 活性以吸引下游 golgin蛋白 Imh1p 坐落在反式高基氏體網 (trans-Golgi network) 上。另外， Arl1p 亦能與 Arf 的交換蛋白 (GEF) Gea2p 以及磷脂轉為蛋白 (flippase) Drs2p 形成 complex 後影響高爾基氏體磷脂組成以調控下游蛋白的募集與否。然而，關於 golgin 蛋白 Imh1p 在細胞中的調控機制和功能仍能不是很清楚。過去研究中已指出 Ras-like 小分子 GTP 結合蛋白 (Rab) Ypt6p主要參與核內體 (endosome) 和高爾基氏體之間的retrograde transport，並推論 Ypt6p 與 Arl1p 的功能性具有重疊性。因此，本研究藉由在剔除Ypt6p 的酵母菌細胞中探討 Arl1p 和 Imh1p 在細胞內的角色。我們發現 Imh1p 為主要 Arl1p 下游作用蛋白抑制在缺失 Ypt6p 細胞的囊泡運輸缺陷，且其需要兩個完全一致且正確配對的 dimerization 才具有功能性。總而言之，我們發現 Arl1p-Imh1p pathway 於囊泡運輸中的重要性以及 Imh1p 的功能調控機制。

關鍵字

ARLs ； Arl1p ； Imh1p ； 反式高爾基氏網 ； 囊泡運輸

並列摘要

ARF-like proteins (ARL) are members of ADP-ribosylation factors (ARF) family of small guanine-nucleotide-binding proteins. Arl1p is known to regulate multiple trafficking pathways at the Golgi. We previously demonstrated that Syt1p, a guanine exchange factor (GEF) for Arl1p, can regulate Arl1p activity and recruit its downstream effector golgin protein Imh1p to the trans-Golgi network (TGN). We also reported that Arf-GEF Gea2p and flippase Drs2p interact with Arl1p to regulate the recruitment of Imh1p to the TGN. However, the biological function of golgin protein Imh1p is still unclear. In this study, we investigate roles of Arl1p and golgin protein Imh1p in vesicle traffic pathway in YPT6-deleted yeast, which lacks a Rab GTPase for the retrograde pathway from the endosome to the TGN. Through ypt6-dependent growth inhibition phenotype, we found that Imh1p is major Arl1p downstream effector involving in Arl1p suppression without Drs2p and Gea2p participation. Furthermore, proper dimerization dynamic and homo-pairing of Imh1p dimer are both required for Imh1p normal function. Together, we demonstrated importance of Arl1p-Imh1p pathway in endosome-to-Golgi traffic, Golgi integrity and regulation mechanism of Imh1p in this study.

並列關鍵字

ARLs ； Arl1p ； Imh1p ； trans-Golgi network (TGN) ； membrane traffic

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