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Identification of a phosphoinositide binding motif that mediates activation of mammalian and yeast phospholipase D isoenzymes
Author(s) -
Sciorra Vicki A.,
Rudge Simon A.,
Prestwich Glenn D.,
Frohman Michael A.,
Engebrecht JoAnne,
Morris Andrew J.
Publication year - 1999
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/18.21.5911
Subject(s) - biology , library science , biological sciences , computational biology , computer science
Phosphoinositides are both substrates for second messenger‐generating enzymes and spatially localized membrane signals that mediate vital steps in signal transduction, cytoskeletal regulation and membrane trafficking. Phosphatidylcholine‐specific phospholipase D (PLD) activity is stimulated by phosphoinositides, but the mechanism and physiological requirement for such stimulation to promote PLD‐dependent cellular processes is not known. To address these issues, we have identified a site at which phosphoinositides interact with PLD and have assessed the role of this region in PLD function. This interacting motif contains critical basic amino acid residues that are required for stimulation of PLD activity by phosphoinositides. Although PLD alleles mutated at this site fail to bind to phosphoinositides in vitro , they are membrane‐associated and properly localized within the cell but are inactive against cellular lipid substrates. Analogous mutations of this site in yeast PLD, Spo14p, result in enzymes that localize normally, but with catalytic activity that has dramatically reduced responsiveness to phosphoinositides. The level of responsiveness to phosphoinositides in vitro correlated with the ability of PLD to function in vivo . Taken together, these results provide the first evidence that phosphoinositide regulation of PLD activity observed in vitro is physiologically important in cellular processes in vivo including membrane trafficking and secretion.