Molecular mechanisms, biological actions, and neuropharmacology of the growth-associated protein GAP-43

Abstract

GAP-43 is an intracellular growth-associated protein that appears to assist neuronal pathfinding and branching during development and regeneration, and may contribute to presynaptic membrane changes in the adult, leading to the phenomena of neurotransmitter release, endocytosis and synaptic vesicle recycling, long-term potentiation, spatial memory formation, and learning. GAP-43 becomes bound via palmitoylation and the presence of three basic residues to membranes of the early secretory pathway. It is then sorted onto vesicles at the late secretory pathway for fast axonal transport to the growth cone or presynaptic plasma membrane. The palmitate chains do not serve as permanent membrane anchors for GAP-43, because at steady-state most of the GAP-43 in a cell is membrane-bound but is not palmitoylated. Filopodial extension and branching take place when GAP-43 is phosphorylated at Ser-41 by protein kinase C, and this occurs following neurotrophin binding and the activation of numerous small GTPases. GAP-43 has been proposed to cluster the acidic phospholipid phosphatidylinositol 4,5-bisphosphate in plasma membrane rafts. Following GAP-43 phosphorylation, this phospholipid is released to promote local actin filament-membrane attachment. The phosphorylation also releases GAP-43 from calmodulin. The released GAP-43 may then act as a lateral stabilizer of actin filaments. N-terminal fragments of GAP-43, containing 10-20 amino acids, will activate heterotrimeric G proteins, direct GAP-43 to the membrane and lipid rafts, and cause the formation of filopodia, possibly by causing a change in membrane tension. This review will focus on new information regarding GAP-43, including its binding to membranes and its incorporation into lipid rafts, its mechanism of action, and how it affects and is affected by extracellular agents.

Keywords: growth cones; neuromodulin; neurotransmitters; neurotrophins; synaptic plasticity.

Fig. (1)

Summary of some proposed mechanisms involved in filopodial formation. These pathways are discussed in detail in the text. The axonal growth cone plasma membrane is shown protruding as a filopodium begins to form. The extracellular matrix protein laminin is shown to the left, near the transmembrane cell adhesion protein, L1. The three actin-binding proteins, gelsolin, profilin, and cofilin, are shown near the F-actin filament, where they have promoted filament dynamics. At this point they are not inactivated by being bound at the plasma membrane to the acidic phospholipid phosphatidylinositol 4,5-bisphosphate, which is designated PI (4,5) P₂ in the text and PIP2 in the diagram, because prior to local stimulation PIP2 is bound to GAP-43, as shown. Upon local stimulation, which in this case is the binding of nerve growth factor (NGF) to its high affinity receptor trk A, as shown at right, a series of events are set into motion. The Rho-GTPase activating protein Grit activates the Rho GTPase cdc42, which promotes filopodial formation. The small GTPase RalA, which along with RalB promotes filopodial formation and neurite branching, activates the exocyst complex, which in turn activates protein kinase C (PKC), located in the center of the diagram. The exocyst also activates cdc42. RalA causes the actin-binding protein filamin to join the F-actin filament, which also aids in the extension of the filopodium. RalB, on the right, activates phospholipase D (PLD), which in turn cleaves the phospholipid phosphatidylcholine (PC) to phosphatidic acid (PA), which is converted to diacylglycerol (DAG). The DAG then stimulates protein kinase C. The activated protein kinase C phosphorylates GAP-43 at Ser-41, as indicated by the arrow, which releases phosphorylated GAP-43 from PIP2 and allows the GAP-43 to potentially act as a lateral stabilizer of the F-actin, shown to the left of GAP-43. The released PIP2 diffuses in the membrane, as shown by the arrow. It then promotes the adhesion of the F-actin with the membrane, and by so doing aids in the formation of the filopodium. This effect also involves the activation of the Wiskott-Aldrich syndrome protein (WASP) by cdc42 and PIP2 and by the recruitment of the Arp2/3 complex (not shown) to the site. WASP binds directly to PIP2, actin filaments, and cdc42. For simplicity, the binding of GAP-43 to calmodulin is not shown, but in the unstimulated cell it would occur similarly to what is shown for the binding of GAP-43 to PIP2. Upon phosphorylation by protein kinase C, GAP-43 would be released from calmodulin, as it is from PIP2, possibly for interaction with F-actin. Unphosphorylated GAP-43, if not bound to PIP2, would act as a capping protein for the actin filament and would oppose filopodial extension. The F-actin shown is part of the membrane skeleton and is also called cell cortex actin. It is possible that GAP-43 is palmitoylated while the above interactions are taking place.