Characterization of the Role of Rab18 in Mediating LD-ER Contact and LD Growth.

Abstract

Lipid droplets (LDs) are dynamic cellular organelles found in most eukaryotic cells. Lipid incorporation from endoplasmic reticulum (ER) to LD is important in controlling LD growth and intracellular lipid homeostasis. However, the molecular link that mediates ER and LD cross talk remains elusive. Here, we describe the methodology used to characterize the function of Rab18 in regulating LD homeostasis and LD-ER contact. First, we focus on the quantitative assay used to measure intracellular LDs morphological changes. This is followed by a detailed description of the use of the APEX-label technology in combination with electron microscope (EM) to visualize ER-LD contact sites. These assays are valuable for the investigation of LD-associated proteins such as Rab18 in establishing membrane contact sites between LDs and other subcellular organelles.