Increased breakdown of phosphatidylinositol 4,5-bisphosphate is not an initiating factor for actin assembly in human neutrophils.
The present study examined the possible role of increased phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) breakdown in the regulation of actin assembly in human neutrophils. Tetracaine, a local anesthetic, was used since it has recently been proposed to inhibit the phosphorylation of phosphatidylinositol 4-phosphate to form PtdIns(4,5)P2. Surprisingly, it was found that incubation with tetracaine alone increased the breakdown of PtdIns(4,5)P2, measured as total inositol trisphosphate formation. This occurred without any rise above basal in the cellular content of filamentous actin. However, in the presence of formylmethionyl-leucyl-phenylalanine (fMet-Leu-Phe), tetracaine potentiated the chemotactic-induced increase of both inositol trisphosphate formation and actin polymerization. To further explore the relationship between increased PtdIns(4,5)P2 breakdown and actin polymerization, the activity of phospholipase C was depressed by lowering the cytosolic free calcium ion level or by incubating the cells with ionomycin. In these cells, fMet-Leu-Phe stimulation still raised the cellular content of filamentous actin to a level similar to levels in nontreated cells, despite the absence of PtdIns(4,5)P2 hydrolysis. Consequently, increased breakdown of PtdIns(4,5)P2 alone is not enough to initiate actin polymerization, nor is the polymerization of actin dependent on an increased PtdIns(4,5)P2 breakdown. However, we cannot exclude the possibility that increased turnover of phosphoinositides might act as a modulator of actin assembly.
Bengtsson
T
T
Department of Medical Microbiology, University of Linköping, Sweden.
Rundquist
I
I
Stendahl
O
O
Wymann
M P
MP
Andersson
T
T
D000199:Actins / Q000096:biosynthesis / Q000097:blood*; D000634:Aminoquinolines / Q000494:pharmacology; D005456:Fluorescent Dyes; D006801:Humans; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate; D007295:Inositol Phosphates / Q000096:biosynthesis / Q000097:blood; D007700:Kinetics; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols / Q000097:blood* / Q000502:physiology; D013748:Tetracaine / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000097:blood
D000199:Actins; D000634:Aminoquinolines; D005456:Fluorescent Dyes; D007295:Inositol Phosphates; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D013748:Tetracaine; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015544:Inositol 1,4,5-Trisphosphate; D010738:Type C Phospholipases; C035739:Quin2