Phospholipase D2 Modulates the Secretory Pathway in RBL-2H3 Mast Cells.
Phospholipase D (PLD) hydrolyses phosphatidylcholine to produce phosphatidic acid (PA) and choline. It has two isoforms, PLD1 and PLD2, which are differentially expressed depending on the cell type. In mast cells it plays an important role in signal transduction. The aim of the present study was to clarify the role of PLD2 in the secretory pathway. RBL-2H3 cells, a mast cell line, transfected to overexpress catalytically active (PLD2CA) and inactive (PLD2CI) forms of PLD2 were used. Previous observations showed that the Golgi complex was well organized in CA cells, but was disorganized and dispersed in CI cells. Furthermore, in CI cells, the microtubule organizing center was difficult to identify and the microtubules were disorganized. These previous observations demonstrated that PLD2 is important for maintaining the morphology and organization of the Golgi complex. To further understand the role of PLD2 in secretory and vesicular trafficking, the role of PLD2 in the secretory process was investigated. Incorporation of sialic acid was used to follow the synthesis and transport of glycoconjugates in the cell lines. The modified sialic acid was subsequently detected by labeling with a fluorophore or biotin to visualize the localization of the molecule after a pulse-chase for various times. Glycoconjugate trafficking was slower in the CI cells and labeled glycans took longer to reach the plasma membrane. Furthermore, in CI cells sialic acid glycans remained at the plasma membrane for longer periods of time compared to RBL-2H3 cells. These results suggest that PLD2 activity plays an important role in regulating glycoconjugate trafficking in mast cells.
Department of Cell and Molecular Biology and Pathogenic Bioagents, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Claudia Maria Meirelles
CM
Marchini-Alves
Department of Cell and Molecular Biology and Pathogenic Bioagents, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Valeria Cintra
VC
Barbosa Lorenzi
Department of Cell and Molecular Biology and Pathogenic Bioagents, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Elaine Zayas Marcelino
EZ
da Silva
Department of Cell and Molecular Biology and Pathogenic Bioagents, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Vivian Marino
VM
Mazucato
Department of Cell and Molecular Biology and Pathogenic Bioagents, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Maria Celia
MC
Jamur
Department of Cell and Molecular Biology and Pathogenic Bioagents, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Constance
C
Oliver
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002460:Cell Line; D008407:Mast Cells / Q000201:enzymology* / Q000378:metabolism*; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D051381:Rats; D055571:Secretory Pathway / Q000502:physiology
C105606:phospholipase D2; D010739:Phospholipase D; D019158:N-Acetylneuraminic Acid