A possible role for mono (ADP-ribosyl) transferase in the signalling pathway mediating neutrophil chemotaxis
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J R Allport1, L E Donnelly, P Kefalas
1Department of Clinical Pharmacology, Royal Postgraduate Medical School, London, UK.
Abstract:
1. Mono(ADP-ribosyl)transferase activity has been identified on the external surface of human polymorphonuclear neutrophil leucocytes (PMNs). The enzyme is released from the plasma membrane by phosphoinositide-specific phospholipase C, suggesting a glycosylphosphatidylinositol (GPI) linkage of the enzyme to the plasma membrane. Partial sequence of cDNA encoding the enzyme suggests that it is identical to the GPI-linked mono(ADP-ribosyl)-transferase identified previously on human skeletal muscle. 2. A panel of inhibitors of mono(ADP-ribosyl)transferase (including vitamins K1 and K3, novobiocin and nicotinamide) showed a rank order of inhibitory potency similar to that described for other mono(ADP-ribosyl)transferases. Furthermore, the mono(ADP-ribosyl)ation of agmatine was inhibited also by diethylamino (benzylidineamino)guanidine (DEA-BAG), another substrate of the enzyme related structurally to arginine. 3. There was a close linear correlation between the IC50 values for inhibition of mono(ADP-ribosyl)ation of agmatine by DEA-BAG or the enzyme inhibitors and their IC50 values for inhibition of receptor-dependent polymerization of cytoskeletal actin and chemotaxis. 4. These results suggest a role for mono(ADP-ribosyl)transferase in the transduction pathway involved in receptor-dependent re-alignment of the cytoskeleton during neutrophil chemotaxis.
Insights
Mono(ADP-ribosyl)transferase on neutrophil surfaces is linked to the plasma membrane via GPI. This enzyme is crucial for cytoskeletal re-alignment during neutrophil chemotaxis.
Area of Science:
- Biochemistry
- Cell Biology
- Immunology
Background:
- Mono(ADP-ribosyl)transferase (MART) activity is present on the outer surface of human polymorphonuclear neutrophil leucocytes (PMNs).
- The enzyme's release by phosphoinositide-specific phospholipase C suggests a glycosylphosphatidylinositol (GPI) linkage to the plasma membrane.
- Partial cDNA sequencing indicates this PMN enzyme is identical to a previously identified GPI-linked MART in human skeletal muscle.
Purpose of the Study:
- To investigate the role of PMN-associated mono(ADP-ribosyl)transferase in neutrophil function.
- To characterize the enzyme's activity and its relationship with cytoskeletal dynamics during chemotaxis.
Main Methods:
- Enzyme activity assays using various inhibitors, including vitamins K1/K3, novobiocin, nicotinamide, and DEA-BAG.
- Measurement of IC50 values for enzyme inhibition and correlation with cytoskeletal actin polymerization and chemotaxis.
- Analysis of enzyme linkage to the plasma membrane using phospholipase C.
Main Results:
- A panel of MART inhibitors demonstrated similar potency ranks as observed for other MART enzymes.
- The mono-ADP-ribosylation of agmatine was inhibited by DEA-BAG, an arginine-like substrate.
- A strong linear correlation was found between IC50 values for MART inhibition and inhibition of actin polymerization and chemotaxis.
Conclusions:
- Mono(ADP-ribosyl)transferase plays a significant role in the signal transduction pathway governing receptor-dependent cytoskeletal re-alignment.
- The enzyme is involved in regulating neutrophil chemotaxis through modulation of the actin cytoskeleton.