Palmitoleate is a mitogen, formed upon stimulation with growth factors, and converted to palmitoleoyl-phosphatidylinositol.
Controversial correlations between biological activity and concentration of the novel lipokine palmitoleate (9Z-hexadecenoate, 16:1) might depend on the formation of an active 16:1 metabolite. For its identification, we analyzed the glycerophospholipid composition of mouse Swiss 3T3 fibroblasts in response to 16:1 using LC-MS/MS. 16:1 was either supplemented to the cell culture medium or endogenously formed when cells were stimulated with insulin or growth factors as suggested by the enhanced mRNA expression of 16:1-biosynthetic enzymes. The proportion of 1-acyl-2-16:1-sn-phosphatidylinositol (16:1-PI) was time-dependently and specifically increased relative to other glycerophospholipids under both conditions and correlated with the proliferation of fatty acid (16:1, palmitate, oleate, or arachidonate)-supplemented cells. Accordingly, cell proliferation was impaired by blocking 16:1 biosynthesis using the selective stearoyl-CoA desaturase-1 inhibitor CAY10566 and restored by supplementation of 16:1. The accumulation of 16:1-PI occurred throughout cellular compartments and within diverse mouse cell lines (Swiss 3T3, NIH-3T3, and 3T3-L1 cells). To elucidate further whether 16:1-PI is formed through the de novo or remodeling pathway of PI biosynthesis, phosphatidate levels and lyso-PI-acyltransferase activities were analyzed as respective markers. The proportion of 16:1-phosphatidate was significantly increased by insulin and growth factors, whereas lyso-PI-acyltransferases showed negligible activity for 16:1-coenzyme A. The relevance of the de novo pathway for 16:1-PI biosynthesis is supported further by the comparable incorporation rate of deuterium-labeled 16:1 and tritium-labeled inositol into PI for growth factor-stimulated cells. In conclusion, we identified 16:1 or 16:1-PI as mitogen whose biosynthesis is induced by growth factors.
Department of Biochemistry and Molecular Biology, Faculty of Medicine, The University of Tokyo, Tokyo 113-0033, Japan.
Andreas
A
Koeberle
Hideo
H
Shindou
Takeshi
T
Harayama
Takao
T
Shimizu
D016475:3T3 Cells; D000818:Animals; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology*; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology*; D051379:Mice; D010716:Phosphatidylinositols / Q000494:pharmacology*; D016133:Polymerase Chain Reaction
D005229:Fatty Acids, Monounsaturated; D036341:Intercellular Signaling Peptides and Proteins; D010716:Phosphatidylinositols; C008757:palmitoleic acid