Submit Data |  Help |  Video Tutorials |  News |  Publications |  FTP Download |  REST API |  Citing RGD |  Contact   

Mechanisms of regulation of phospholipase D1 by protein kinase Calpha.

Authors: Hu, T  Exton, JH 
Citation: Hu T and Exton JH, J Biol Chem 2003 Jan 24;278(4):2348-55.
Pubmed: (View Article at PubMed) PMID:12431982
DOI: Full-text: DOI:10.1074/jbc.M210093200

It has been suggested that protein-protein interaction is important for protein kinase C (PKC) alpha to activate phospholipase D1 (PLD1). To determine the one or more sites on PKCalpha that are involved in binding to PLD1, fragments containing the regulatory domain, catalytic domain, and C1-C3 domain of PKCalpha were constructed and shown to be functional, but they all failed to bind and activate PLD1 in vivo and in vitro. A C-terminal 23-amino acid (aa) deletion mutant of PKCalpha was also found to be inactive. To define the binding/activation site(s) in the C terminus of PKCalpha, 1- to 11-aa deletion mutants were made in this terminus. Deletion of up to 9 aa did not alter the ability of PKCalpha to bind and activate PLDl, whereas a 10-aa deletion was inactive. The residue at position 10 was Phe(663). Mutations of this residue (F663D and F663A) caused loss of binding, activation, and phosphorylation of PLD1, indicating that Phe(663) is essential for these activities. Time course experiments showed that the activation of PLD1 by PMA was much faster than its phosphorylation, and its activity decreased as phosphorylation increased with time. Staurosporine, a PKC inhibitor, completely inhibited PLD1 phosphorylation in response to 4beta-phorbol 12-myristate 13-acetate PMA and blocked the later decrease in PLD activity. The same results were found with the D481E mutant of PKCalpha, which is unable to phosphorylate PLD1. These results indicate that neither the regulatory nor catalytic domains of PKCalpha alone can bind to or activate PLD1 and that a residue in the C terminus of PKCalpha (Phe(663)) is required for these effects. The initial activation of PLD1 by PMA is highly correlated with the binding of PKCalpha. Although PKCalpha can phosphorylate PLD1, this is a relatively slow process and is associated with inactivation of the enzyme.

Annotation

Gene Ontology Annotations
Objects Annotated

Additional Information

 
RGD Object Information
RGD ID: 729596
Created: 2003-11-26
Species: All species
Last Modified: 2004-05-25
Status: ACTIVE



NHLBI Logo

RGD is funded by grant HL64541 from the National Heart, Lung, and Blood Institute on behalf of the NIH.