Division of Molecular Interaction, Institute for Genetic Medicine, Hokkaido University Graduate School of Life Science, Sapporo, Hokkaido, Japan.
Department of Gastroenterological Surgery II, Hokkaido University Faculty of Medicine, Sapporo, Hokkaido, Japan.
PLoS One. 2020 Jul 30;15(7):e0236520. doi: 10.1371/journal.pone.0236520. eCollection 2020.
In eukaryotic cells, phospholipid flippases translocate phospholipids from the exoplasmic to the cytoplasmic leaflet of the lipid bilayer. Budding yeast contains five flippases, of which Cdc50p-Drs2p and Neo1p are primarily involved in membrane trafficking in endosomes and Golgi membranes. The ANY1/CFS1 gene was identified as a suppressor of growth defects in the neo1Δ and cdc50Δ mutants. Cfs1p is a membrane protein of the PQ-loop family and is localized to endosomal/Golgi membranes, but its relationship to phospholipid asymmetry remains unknown. The neo1Δ cfs1Δ mutant appears to function normally in membrane trafficking but may function abnormally in the regulation of phospholipid asymmetry. To identify a gene that is functionally relevant to NEO1 and CFS1, we isolated a mutation that is synthetically lethal with neo1Δ cfs1Δ and identified ERD1. Erd1p is a Golgi membrane protein that is involved in the transport of phosphate (Pi) from the Golgi lumen to the cytoplasm. The Neo1p-depleted cfs1Δ erd1Δ mutant accumulated plasma membrane proteins in the Golgi, perhaps due to a lack of phosphatidylinositol 4-phosphate. The Neo1p-depleted cfs1Δ erd1Δ mutant also exhibited abnormal structure of the endoplasmic reticulum (ER) and induced an unfolded protein response, likely due to defects in the retrieval pathway from the cis-Golgi region to the ER. Genetic analyses suggest that accumulation of Pi in the Golgi lumen is responsible for defects in Golgi functions in the Neo1p-depleted cfs1Δ erd1Δ mutant. Thus, the luminal ionic environment is functionally relevant to phospholipid asymmetry. Our results suggest that flippase-mediated phospholipid redistribution and luminal Pi concentration coordinately regulate Golgi membrane functions.
在真核细胞中，磷脂翻转酶将磷脂从质膜的外叶层转移到细胞质叶层。酿酒酵母含有五种翻转酶，其中 Cdc50p-Drs2p 和 Neo1p 主要参与内体和高尔基体膜的膜运输。ANY1/CFS1 基因被鉴定为 neo1Δ 和 cdc50Δ 突变体生长缺陷的抑制因子。Cfs1p 是 PQ-环家族的膜蛋白，定位于内体/高尔基体膜，但它与磷脂不对称性的关系尚不清楚。neo1Δ cfs1Δ 突变体在膜运输中似乎正常发挥功能，但在磷脂不对称性的调节中可能功能异常。为了鉴定与 NEO1 和 CFS1 功能相关的基因，我们分离了一个与 neo1Δ cfs1Δ 突变体合成致死的突变体，并鉴定出 ERD1。Erd1p 是一种高尔基体膜蛋白，参与从高尔基体腔到细胞质的磷酸盐（Pi）运输。在 Neo1p 耗尽的 cfs1Δ erd1Δ 突变体中，质膜蛋白在高尔基体中积累，这可能是由于缺乏磷脂酰肌醇 4-磷酸。Neo1p 耗尽的 cfs1Δ erd1Δ 突变体还表现出内质网 (ER) 的异常结构，并诱导未折叠蛋白反应，这可能是由于从顺式高尔基体区域到 ER 的回收途径缺陷所致。遗传分析表明，高尔基体腔中 Pi 的积累是导致 Neo1p 耗尽的 cfs1Δ erd1Δ 突变体中高尔基体功能缺陷的原因。因此，高尔基体腔中的离子环境与磷脂不对称性的功能相关。我们的结果表明，翻转酶介导的磷脂再分配和腔中 Pi 浓度共同调节高尔基体膜功能。
