Arf1-dependent PLD1 is localized to oleic acid-induced lipid droplets in NIH3T3 cells.
Phospholipase D (PLD) is known to play a role in vesicle transport through the hydrolysis of phosphatidylcholine (PC) to produce the bioactive lipid, phosphatidic acid. Lipid droplets (LDs) are surrounded by a monolayer of phospholipids, including PC and its lyso derivative, and exhibit a number of signaling proteins. Our recent report suggests that the association of adipose differentiation-related protein (ADRP) to LDs is regulated by an ADP-ribosylation factor 1 (Arf1)-dependent mechanism. In the present study, we found an increase in PLD activity accompanied with LD formation in oleic acid-treated NIH3T3 cells. Brefeldin A, an inhibitor of ARF-GEFs, suppressed both PLD activation and LD formation in oleic acid-treated cells. PLD1, but not PLD2, was found to exist in LDs by immunocytochemical analysis. Furthermore, co-existence of PLD1, Arf1, and ADRP was observed in the LD-enriched subcellular fractions obtained from oleic acid-treated NIH3T3 cells by Western blot analysis. PLD1 activity in the LD-enriched fractions was stimulated by exogenously added Arf1. Although LDs were induced in either PLD1- or PLD2-overexpressing CHO cells by oleic acid treatment, the stimulation of PLD activity was observed only in PLD1-CHO cells. Taken together, the data suggest that the activation of Arf1-dependent PLD1 occurs in LDs and may be involved in their physiological function.
Nakamura
Noriko
N
Department of Anatomy and Molecular Cell Biology, Nagoya University, Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan. nakamur1@niehs.nih.gov
Banno
Yoshiko
Y
Tamiya-Koizumi
Keiko
K
D020823:ADP-Ribosylation Factor 1 / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D022162:Cytoplasmic Vesicles / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D019301:Oleic Acid / Q000494:pharmacology*; D000070780:Perilipin-2; D010739:Phospholipase D / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism*; D021381:Protein Transport; D013347:Subcellular Fractions
D008565:Membrane Proteins; D000070780:Perilipin-2; D010743:Phospholipids; C000602983:Plin2 protein, mouse; D019301:Oleic Acid; D010739:Phospholipase D; C106801:phospholipase D1; D020823:ADP-Ribosylation Factor 1