Expression and regulation of phospholipase D in the human keratinocyte cell line HaCaT

FEBS Letters

Höer, Ariane²  Riebeling, Christian¹  Wieder, Thomas¹  Orfanos, Constantin E¹  Scholz, Frank-Rüdiger¹  Müller-Wieprecht, Verena¹  Alexander, Christian¹  Geilen, Christoph C¹ 

[1] Department of Dermatology, University Medical Center Benjamin Franklin, The Free University of Berlin, Hindenburgdamm 30, D-12200 Berlin, Germany;Department of Pharmacology and Toxicology, University Medical Center Benjamin Franklin, The Free University of Berlin, Hindenburgdamm 30, D-12200 Berlin, Germany

关键词: Phospholipase D;    Signal transduction;    HaCaT cell;    Peptide-specific antibody;    ARF1;    ADP-ribosylation factor 1;    BCA;    bicinchoninic acid;    BSA;    bovine serum albumin;    DPPC;    dipalmitoylphosphatidylcholine;    EGTA;    ethylene glycol-bis[β-aminoethylether]-N;    N;    N′;    N′-tetraacetic acid;    GTPγS;    guanosine 5′-O-(3-thiotriphosphate);    Na-HEPES;    N-[2-hydroxyethyl]piperazine-N′-[2-ethanesulfonic acid] sodium salt;    MgATP;    adenosine triphosphate magnesium salt;    PC;    phosphatidylcholine;    PE;    phosphatidylethanolamine;    PEt;    phosphatidylethanol;    PIP2;    1-phosphatidyinositol-4;    5-bisphosphate;    PKC;    protein kinase C;    PLD;    phospholipase D;    PMSF;    phenylmethylsulfonyl fluoride;    SDS-PAGE;    sodium dodecylsulfate-polyacrylamide gel electrophoresis;    TPA;    12-O-tetradecanoyl phorbol-13-acetate;   

DOI  :  10.1016/S0014-5793(98)00227-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The generation of lipid second messengers via phosphatidylcholine (PC)-specific phospholipase D (PLD) has emerged as an important step leading to transduction of extracellular signals. In the present investigation the expression of human cytosolic PLD isoenzymes in the immortalized human keratinocyte cell line HaCat was determined. At the mRNA level we found the expression of hPLD1b and for the first time in human cells also the expression of hPLD2. For further analysis of enzyme expression at the protein level, hPLD1 peptide fragments were synthesized and specific antibodies were generated (rabbit) to be used for detection of hPLD1 in Western blot experiments. Furthermore, small G-proteins were found to be involved in the regulation of PLD activity in HaCaT cells using the guanine nucleotide analogue GTPγS.

【 授权许可】

Unknown