Regulation of phospholipase C delta1 by sphingosine.
Sphingosine, which is on the pathway of sphingomyelin degradation, activates phospholipase C (PLC) delta1 moderately. In the liposome assay effect of sphingosine on PLC delta1 activity depends on KCl concentration. Stimulation of PLC delta1 by sphingosine increased as the KCl concentration is increased from 0 to 100 mM, and then diminished with the increasing KCl. In the liposome assay sphingosine diminishes inhibition of PLC delta1 by sphingomyelin. To determine the domain of PLC delta1 which interacts with sphingosine active proteolytic fragments of PLC delta1 were generated by trypsin digestion of the native enzyme. Sphingosine affects the activity of PLC delta1 fragment which lacked the amino-terminal domain (first 60 amino acids) but not the active fragment that has cleaved the domain spanning the X and Y region of PLC delta1. These observations indicate that for interaction of sphingosine with PLC delta1 intact domain that span regions of conservation, designated as X and Y is necessary. When the activity of PLC delta1 was assayed with PIP2 in the erythrocyte membrane as substrate, sphingosine strongly inhibited PLC delta1. The other homolog of sphingosine 4-hydroxysphinganine (phytosphingosine) inhibited PLC delta1 to much lesser extent. The activity of PLC delta1 was inhibited by 68% and 22% in the presence of 20 microM sphingosine and phytosphingosine, respectively. This inhibition was completely abolished by deoxycholate at a concentration of 1.5 mM. These observations suggest that sphingosine may regulate activity of PLC delta1 in the cell.
Matecki
A
A
Department of Clinical Biochemistry, Medical University of Gdansk, Poland.
Pawelczyk
T
T
D001667:Binding, Competitive; D003902:Detergents; D004789:Enzyme Activation; D004910:Erythrocyte Membrane / Q000201:enzymology; D006801:Humans; D008081:Liposomes; D010446:Peptide Fragments / Q000032:analysis; D010743:Phospholipids / Q000378:metabolism; D013109:Sphingomyelins / Q000378:metabolism; D013110:Sphingosine / Q000494:pharmacology*; D014357:Trypsin; D010738:Type C Phospholipases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*
D003902:Detergents; D008081:Liposomes; D010446:Peptide Fragments; D010743:Phospholipids; D013109:Sphingomyelins; D010738:Type C Phospholipases; D014357:Trypsin; D013110:Sphingosine