Quantitative live-cell PALM reveals nanoscopic Faa4 redistributions and dynamics on lipid droplets during metabolic transitions of yeast.
Quantitative live-cell PALM reveals nanoscopic Faa4 redistributions and dynamics on lipid droplets during metabolic transitions of yeast.
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DOI：
10.1091/mbc.e20-11-0695
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发表时间：
2021-08-15
影响因子：
3.3
通讯作者：
Puchner EM
中科院分区：
文献类型：
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作者：
Adhikari S;Moscatelli J;Puchner EM
脂滴是一种动态的脂质储存和体内平衡的细胞器。细胞通过调节LD的空间分布和LD生长和周转所需的酶来响应代谢变化。ld的小尺寸妨碍了用常规荧光显微镜观察它们相关的酶密度和动力学。本研究采用定量光激活定位显微镜技术，以单分子灵敏度和30 nm分辨率研究了酵母活细胞ld上脂肪酸激活酶Faa4的密度。在日志期，ld与内质网（ER）共定位，它们的出现和扩张由观察到的最高Faa4密度介导。在过渡到固定相期间，表面积增加2倍的ld转移到液泡表面和管腔，并表现出Faa4密度增加2.5倍。LD上Faa4密度的增加进一步表明，它在LD扩张中的作用是由其表达水平增加约5倍引起的，并且与外源FA链长度有关。当被更新的培养基诱导脂肪分解时，Faa4通过ER-和脂质吞噬穿梭到液泡，在那里它可能激活FAs进行膜扩张并降解Faa4，使其细胞丰度恢复到log期的水平。
Lipid droplets (LDs) are dynamic organelles for lipid storage and homeostasis. Cells respond to metabolic changes by regulating the spatial distribution of LDs and enzymes required for LD growth and turnover. The small size of LDs precludes the observation of their associated enzyme densities and dynamics with conventional fluorescence microscopy. Here we employ quantitative photo-activated localization microscopy to study the density of the fatty acid (FA) activating enzyme Faa4 on LDs in live yeast cells with single-molecule sensitivity and 30 nm resolution. During the log phase LDs colocalize with the endoplasmic reticulum (ER) where their emergence and expansion are mediated by the highest observed Faa4 densities. During transition to the stationary phase, LDs with a ∼2-fold increased surface area translocate to the vacuolar surface and lumen and exhibit a ∼2.5-fold increase in Faa4 density. The increased Faa4 density on LDs further suggests its role in LD expansion, is caused by its ∼5-fold increased expression level, and is specific to exogenous FA chain-lengths. When lipolysis is induced by refreshed medium, Faa4 shuttles through ER- and lipophagy to the vacuole, where it may activate FAs for membrane expansion and degrade Faa4 to reset its cellular abundance to levels in the log phase.