Regulated localization of Rab18 to lipid droplets: effects of lipolytic stimulation and inhibition of lipid droplet catabolism.
Rab GTPases are crucial regulators of membrane traffic. Here we have examined a possible association of Rab proteins with lipid droplets (LDs), neutral lipid-containing organelles surrounded by a phospholipid monolayer, also known as lipid bodies, which have been traditionally considered relatively inert storage organelles. Although we found close apposition between LDs and endosomal compartments labeled by expressed Rab5, Rab7, or Rab11 constructs, there was no detectable labeling of the LD surface itself by these Rab proteins. In contrast, GFP-Rab18 localized to LDs and immunoelectron microscopy showed direct association with the monolayer surface. Green fluorescent protein (GFP)-Rab18-labeled LDs underwent oscillatory movements in a localized area as well as sporadic, rapid, saltatory movements both in the periphery of the cell and toward the perinuclear region. In both adipocytes and non-adipocyte cell lines Rab18 localized to a subset of LDs. To gain insights into this specific localization, Rab18 was co-expressed with Cav3DGV, a truncation mutant of caveolin-3 shown to inhibit the catabolism and motility of lipid droplets. GFP-Rab18 and mRFP-Cav3DGV labeled mutually exclusive subpopulations of LDs. Moreover, in 3T3-L1 adipocytes, stimulation of lipolysis increased the localization of Rab18 to LDs, an effect reversed by beta-adrenergic antagonists. These results show that a Rab protein localizes directly to the monolayer surface of LDs. In addition, association with the LD surface was increased following stimulation of lipolysis and inhibited by a caveolin mutant suggesting that recruitment of Rab18 is regulated by the metabolic state of individual LDs.
Martin
Sally
S
Institute for Molecular Bioscience & Centre for Microscopy and Microanalysis, University of Queensland, Brisbane, Queensland 4072, Australia.
Driessen
Kim
K
Nixon
Susan J
SJ
Zerial
Marino
M
Parton
Robert G
RG
D041721:3T3-L1 Cells; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D049452:Green Fluorescent Proteins / Q000378:metabolism; D050356:Lipid Metabolism; D008066:Lipolysis; D051379:Mice; D008854:Microscopy, Electron; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
D017931:DNA Primers; C083879:Rab18 protein, mouse; D049452:Green Fluorescent Proteins; D020691:rab GTP-Binding Proteins