Repulsive axon guidance cues ephrin-A2 and slit3 stop protrusion of the growth cone leading margin concurrently with inhibition of ADF/cofilin and ERM proteins.

Axonal growth cones turn away from repulsive guidance cues. This may start with reduced protrusive motility in the region the growth cone leading margin that is closer to the source of repulsive cue. Using explants of E7 chick temporal retina, we examine the effects of two repulsive guidance cues, ephrin-A2 and slit3, on retinal ganglion cell growth cone protrusive activity, total F-actin, free F-actin barbed ends, and the activities (phosphorylation states) of actin regulatory proteins, ADF/cofilin and ezrin, radixin, moesin (ERM) proteins.

Activation of ezrin/radixin/moesin mediates attractive growth cone guidance through regulation of growth cone actin and adhesion receptors.

The development of a functioning neural network relies on responses of axonal growth cones to molecular guidance cues that are encountered en route to their target tissue. Nerve growth factor (NGF) and neurotrophin-3 serve as attractive cues for chick embryo sensory growth cones in vitro and in vivo, but little is known about the actin-binding proteins necessary to mediate this response. The evolutionarily conserved ezrin/radixin/moesin (ERM) family of proteins can tether actin filaments to the cell membrane when phosphorylated at a conserved threonine residue.

Labeling F-actin barbed ends with rhodamine-actin in permeabilized neuronal growth cones.

The motile tips of growing axons are called growth cones. Growth cones lead navigating axons through developing tissues by interacting with locally expressed molecular guidance cues that bind growth cone receptors and regulate the dynamics and organization of the growth cone cytoskeleton. The main target of these navigational signals is the actin filament meshwork that fills the growth cone periphery and that drives growth cone motility through continual actin polymerization and dynamic remodeling.

Activation of ADF/cofilin mediates attractive growth cone turning toward nerve growth factor and netrin-1.

Proper neural circuitry requires that growth cones, motile tips of extending axons, respond to molecular guidance cues expressed in the developing organism. However, it is unclear how guidance cues modify the cytoskeleton to guide growth cone pathfinding. Here, we show acute treatment with two attractive guidance cues, nerve growth factor (NGF) and netrin-1, for embryonic dorsal root ganglion and temporal retinal neurons, respectively, results in increased growth cone membrane protrusion, actin polymerization, and filamentous actin (F-actin).

Protein synthesis in distal axons is not required for growth cone responses to guidance cues.

Recent evidence suggests that growth cone responses to guidance cues require local protein synthesis. Using chick neurons, we investigated whether protein synthesis is required for growth cones of several types to respond to guidance cues. First, we found that global inhibition of protein synthesis stops axonal elongation after 2 h.