| 盛园园,高关刚,李域琪,金美君,徐 俐.PAT蛋白抑制饥饿诱导的脂滴快速融合[J].,2020,(4):601-607 |
| PAT蛋白抑制饥饿诱导的脂滴快速融合 |
| PAT Proteins Inhibit Starvation-induced Lipid Droplet Coalescence |
| 投稿时间：2019-08-15 修订日期：2019-09-10 |
| DOI：10.13241/j.cnki.pmb.2020.04.001 |
| 中文关键词: 脂滴 快速融合 PAT蛋白 自噬 |
| 英文关键词: Lipid droplet Coalescence PAT proteins Autophagy |
| 基金项目: |
|
| 摘要点击次数: 925 |
| 全文下载次数: 634 |
| 中文摘要: |
| 摘要 目的：脂滴快速融合是增大脂滴直径的方式之一，但其研究相对少。本研究旨在建立脂滴快速融合的细胞模型，以便对其进行深入的生物学研究。方法：本研究使用大鼠肾成纤维细胞系NRK和小鼠前脂肪细胞系3T3-L1两种细胞系，先用油酸诱导细胞内产生大量脂滴，再使用饥饿缓冲液培养细胞，利用显微镜实时观测技术跟踪脂滴动态变化，建立脂滴快速融合的模型。而后在此模型中，加入自噬抑制剂或者以过表达CCT为阳性对照，过表达PAT蛋白（PLIN1、ADRP和TIP47），来探究它们在调控脂滴快速融合方面的功能。结果：饥饿缓冲液处理约3小时可诱导细胞发生脂滴快速融合，其融合速率很快，从脂滴接触到融合完成可发生在20秒内，显然不同于CIDE蛋白调控的缓慢脂滴融合过程。自噬抑制剂可以抑制自噬，但是并没有显著影响脂滴快速融合，说明饥饿诱导的脂滴快速融合不依赖于自噬。另发现，与过表达GFP相比，过表达定位于脂滴的GFP-CCT、GFP-PLIN1、GFP-ADRP或GFP-TIP47均能显著性抑制快速融合导致的脂滴变大的现象。结论：本研究建立了饥饿缓冲液诱导脂滴发生快速融合的细胞模型，并证明PAT蛋白（PLIN1、ADRP、TIP47）能抑制脂滴快速融合。 |
| 英文摘要: |
| ABSTRACT Objective: Lipid droplet coalescence is one of the ways to effectively increase lipid droplet (LD) size. However, compared with other methods, this method is less researched. This study aimed to establish a cell model of LD coalescence for its further biological study. Methods: Two cell lines, namely rat kidney fibroblast cell line NRK and mouse pre-adipocyte cell line 3T3-L1, were cultured with oleic acid (OA) to induce a large number of cytosolic LDs. To establish lipid droplet coalescence model, the cells were cultured with starvation buffer, and the dynamic changes of LDs were tracked by time-lapse observation with confocal microscopy. Then, by adding autophagy inhibitors or overexpressing PAT proteins (PLIN1/ADRP/TIP47) with overexpressing CCT as a positive control, their roles in regulating LD coalescence were studied in this model. Results: Starvation-induced LD coalescence was obviously observed about 3 hours after the starvation buffer treatment. The LD coalescence process was characterized by a fast fusion as we observed the process could be completed within 20 seconds from two LDs contact to their fusion into a large LD. The coalescence process was different from CIDE-mediated LD fusion which was a slow process. Autophagy inhibitors could inhibit autophagy, but did not significantly alter LD coalescence, indicating that starvation-induced LD coalescence did not depend on autophagy. Further, compared with overexpressing GFP, overexpressing LD proteins, including GFP-CCT, GFP-PLIN1, GFP-ADRP, or GFP-TIP47, significantly blocked LD diameter increase caused by starvation-induced LD coalescence. Conclusion: This study established a starvation-induced LD coalescence cell model and demonstrated PAT proteins (PLIN1, ADRP, TIP47) inhibited LD coalescence. |
|
查看全文
查看/发表评论 下载PDF阅读器 |
| 关闭 |
|
|
|
