Ozeki Shintaro, Cheng Jinglei, Tauchi-Sato Kumi, Hatano Naoya, Taniguchi Hisaaki, Fujimoto Toyoshi
Department of Anatomy and Molecular Cell Biology, Graduate School of Medicine, Nagoya University, Showa, Japan.
J Cell Sci. 2005 Jun 15;118(Pt 12):2601-11. doi: 10.1242/jcs.02401. Epub 2005 May 24.
Lipid droplets (LDs) are organelles that store neutral lipids, but their regulatory mechanism is not well understood. In the present study, we identified Rab18 as an LD component of HepG2 cells by proteomic analysis, and confirmed its localization by immunohistochemistry and western blotting. Wild-type and dominant-active Rab18 localized to LDs but the dominant-negative form did not. Endogenous Rab18 coexisted with adipocyte differentiation-related protein (ADRP) in LDs, but the labeling intensity of the two proteins showed clear reciprocity. Consistent with this observation, overexpression of Rab18 induced a decrease in the amounts of ADRP in LDs in HepG2 and BALB/c 3T3 cells. Furthermore, Rab18 overexpression caused close apposition of LDs to membrane cisternae connected to the rough ER. Two other procedures that decrease ADRP, i.e. RNA interference and brefeldin A treatment, induced the same morphological change, indicating that decrease in ADRP was the cause of the LD-ER apposition. In accordance with similar structures found between ER and other organelles, we propose that the ER membrane apposed to LDs should be named the LD-associated membrane, or LAM. The present results suggested that Rab18 regulates LAM formation, which is likely to be involved in mobilizing lipid esters stored in LDs.
脂滴（LDs）是储存中性脂质的细胞器，但其调节机制尚未完全清楚。在本研究中，我们通过蛋白质组学分析鉴定出Rab18是HepG2细胞的一种脂滴成分，并通过免疫组织化学和蛋白质印迹法证实了其定位。野生型和显性活性Rab18定位于脂滴，但显性阴性形式则不然。内源性Rab18与脂肪细胞分化相关蛋白（ADRP）在脂滴中共存，但这两种蛋白的标记强度呈现明显的相互关系。与此观察结果一致，Rab18的过表达导致HepG2和BALB/c 3T3细胞中脂滴内ADRP的量减少。此外，Rab18的过表达使脂滴与连接到粗面内质网的膜池紧密相邻。另外两种降低ADRP的方法，即RNA干扰和布雷菲德菌素A处理，也诱导了相同的形态变化，表明ADRP的减少是脂滴与内质网相邻的原因。根据在内质网和其他细胞器之间发现的类似结构，我们建议将与脂滴相邻的内质网膜命名为脂滴相关膜（LAM）。目前的结果表明，Rab18调节LAM的形成，这可能参与动员储存在脂滴中的脂酯。
