title: Regulation of Macropinocytosis by Diacylglycerol Kinase ζ.
abstract: Macropinosomes arise from the closure of plasma membrane ruffles to bring about the non-selective uptake of nutrients and solutes into cells. The morphological changes underlying ruffle formation and macropinosome biogenesis are driven by actin cytoskeleton rearrangements under the control of the Rho GTPase Rac1. We showed previously that Rac1 is activated by diacylglycerol kinase ζ (DGKζ), which phosphorylates diacylglycerol to yield phosphatidic acid. Here, we show DGKζ is required for optimal macropinocytosis induced by growth factor stimulation of mouse embryonic fibroblasts. Time-lapse imaging of live cells and quantitative analysis revealed DGKζ was associated with membrane ruffles and nascent macropinosomes. Macropinocytosis was attenuated in DGKζ-null cells, as determined by live imaging and vaccinia virus uptake experiments. Moreover, macropinosomes that did form in DGKζ-null cells were smaller than those found in wild type cells. Rescue of this defect required DGKζ catalytic activity, consistent with it also being required for Rac1 activation. A constitutively membrane bound DGKζ mutant substantially increased the size of macropinosomes and potentiated the effect of a constitutively active Rac1 mutant on macropinocytosis. Collectively, our results suggest DGKζ functions in concert with Rac1 to regulate macropinocytosis.
authors.affiliation: Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Rd, Ottawa, ON, K1H 8M5, Canada.|Centre for Neuromuscular Disease, University of Ottawa, Ottawa, ON, Canada.
authors.forename: Ryan
authors.initials: R
authors.lastname: Ard
authors.affiliation: Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Rd, Ottawa, ON, K1H 8M5, Canada.|Centre for Neuromuscular Disease, University of Ottawa, Ottawa, ON, Canada.
authors.forename: Kirk
authors.initials: K
authors.lastname: Mulatz
authors.affiliation: Centre for Innovative Cancer Research, Ottawa Hospital Research Institute, 503 Smyth Road, 3rd floor ORCC, Ottawa, ON, K1H 1C4, Canada.|Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada.
authors.forename: Julia L
authors.initials: JL
authors.lastname: Pomoransky
authors.affiliation: Centre for Neuromuscular Disease, University of Ottawa, Ottawa, ON, Canada.|Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada.|Regenerative Medicine Program, Ottawa Hospital Research Institute, 501 Smyth Road, Ottawa, ON, K1H 8L6, Canada.
authors.forename: Robin J
authors.initials: RJ
authors.lastname: Parks
authors.affiliation: Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Rd, Ottawa, ON, K1H 8M5, Canada.|Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, ON, Canada.
authors.forename: Laura
authors.initials: L
authors.lastname: Trinkle-Mulcahy
authors.affiliation: Centre for Innovative Cancer Research, Ottawa Hospital Research Institute, 503 Smyth Road, 3rd floor ORCC, Ottawa, ON, K1H 1C4, Canada.|Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada.
authors.forename: John C
authors.initials: JC
authors.lastname: Bell
authors.affiliation: Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Rd, Ottawa, ON, K1H 8M5, Canada.|Centre for Neuromuscular Disease, University of Ottawa, Ottawa, ON, Canada.
authors.forename: Stephen H
authors.initials: SH
authors.lastname: Gee
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D019852:Diacylglycerol Kinase / Q000502:physiology*; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D010766:Phosphorylation; D010873:Pinocytosis / Q000502:physiology*; D015398:Signal Transduction; D059008:Time-Lapse Imaging; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
chemical_list: D019852:Diacylglycerol Kinase; C512480:diacylglycerol kinase zeta, mouse; D020830:rac1 GTP-Binding Protein