Frazzled promotes growth cone attachment at the source of a Netrin gradient in the Drosophila visual system.

Axon guidance is proposed to act through a combination of long- and short-range attractive and repulsive cues. The ligand-receptor pair, Netrin (Net) and Frazzled (Fra) (DCC, Deleted in Colorectal Cancer, in vertebrates), is recognized as the prototypical effector of chemoattraction, with roles in both long- and short-range guidance. In the Drosophila visual system, R8 photoreceptor growth cones were shown to require Net-Fra to reach their target, the peak of a Net gradient. Using live imaging, we show, however, that R8 growth cones reach and recognize their target without Net, Fra, or Trim9, a conserved binding partner of Fra, but do not remain attached to it. Thus, despite the graded ligand distribution along the guidance path, Net-Fra is not used for chemoattraction. Based on findings in other systems, we propose that adhesion to substrate-bound Net underlies both long- and short-range Net-Fra-dependent guidance in vivo, thereby eroding the distinction between them

WH2 and proline-rich domains of WASP-family proteins collaborate to accelerate actin filament elongation.

WASP-family proteins are known to promote assembly of branched actin networks by stimulating the filament-nucleating activity of the Arp2/3 complex. Here, we show that WASP-family proteins also function as polymerases that accelerate elongation of uncapped actin filaments. When clustered on a surface, WASP-family proteins can drive branched actin networks to grow much faster than they could by direct incorporation of soluble monomers. This polymerase activity arises from the coordinated action of two regulatory sequences: (i) a WASP homology 2 (WH2) domain that binds actin, and (ii) a proline-rich sequence that binds profilin-actin complexes. In the absence of profilin, WH2 domains are sufficient to accelerate filament elongation, but in the presence of profilin, proline-rich sequences are required to support polymerase activity by (i) bringing polymerization-competent actin monomers in proximity to growing filament ends, and (ii) promoting shuttling of actin monomers from profilin-actin complexes onto nearby WH2 domains. Unoccupied WH2 domains transiently associate with free filament ends, preventing their growth and dynamically tethering the branched actin network to the WASP-family proteins that create it. Collaboration between WH2 and proline-rich sequences thus strikes a balance between filament growth and tethering. Our work expands the number of critical roles that WASP-family proteins play in the assembly of branched actin networks to at least three: (i) promoting dendritic nucleation; (ii) linking actin networks to membranes; and (iii) accelerating filament elongation

In Silico Reconstitution of Actin-Based Symmetry Breaking and Motility

Computational modeling and experimentation in a model system for actin-based force generation explain how actin networks initiate and maintain directional movement

Biochemical Control of Actin-Based Motility: Regulation of Arp2/3 Complex Nucleation by Capping Protein

Capping protein is an integral component of Arp2/3-nucleated actin networks that drive amoeboid motility. Increasing the concentration of capping protein, which caps barbed ends of actin filaments and prevents elongation, increases the rate of actin-based motility in vivo and in vitro. We studied the synergy between capping protein and Arp2/3 using an in vitro actin-based motility system reconstituted from purified proteins. We find that capping protein increases the rate of motility by promoting more frequent filament nucleation by the Arp2/3 complex, and not by increasing the rate of filament elongation as previously suggested. One consequence of this coupling between capping and nucleation is that, while the rate of motility depends strongly on the concentration of capping protein and Arp2/3, the net rate of actin assembly is insensitive to changes in either factor. By reorganizing their architecture, dendritic actin networks harness the same assembly kinetics to drive different rates of motility

Frazzled promotes growth cone attachment at the source of a Netrin gradient in the Drosophila visual system.

Axon guidance is proposed to act through a combination of long- and short-range attractive and repulsive cues. The ligand-receptor pair, Netrin (Net) and Frazzled (Fra) (DCC, Deleted in Colorectal Cancer, in vertebrates), is recognized as the prototypical effector of chemoattraction, with roles in both long- and short-range guidance. In the Drosophila visual system, R8 photoreceptor growth cones were shown to require Net-Fra to reach their target, the peak of a Net gradient. Using live imaging, we show, however, that R8 growth cones reach and recognize their target without Net, Fra, or Trim9, a conserved binding partner of Fra, but do not remain attached to it. Thus, despite the graded ligand distribution along the guidance path, Net-Fra is not used for chemoattraction. Based on findings in other systems, we propose that adhesion to substrate-bound Net underlies both long- and short-range Net-Fra-dependent guidance in vivo, thereby eroding the distinction between them

Developmental neural activity requires neuron-astrocyte interactions.

Developmental neural activity is a common feature of neural circuit assembly. Although glia have established roles in synapse development, the contribution of neuron-glia interactions to developmental activity remains largely unexplored. Here we show that astrocytes are necessary for developmental activity during synaptogenesis in Drosophila. Using wide-field epifluorescence and two-photon imaging, we show that the glia of the central nervous system participate in developmental activity with type-specific patterns of intracellular calcium dynamics. Genetic ablation of astrocytes, but not of cortex or ensheathing glia, leads to severe attenuation of neuronal activity. Similarly, inhibition of neuronal activity results in the loss of astrocyte calcium dynamics. By altering these dynamics, we show that astrocytic calcium cycles can influence neuronal activity but are not necessary per se. Taken together, our results indicate that, in addition to their recognized role in the structural maturation of synapses, astrocytes are also necessary for the function of synapses during development

A discrete neuronal population coordinates brain-wide developmental activity.

In vertebrates, stimulus-independent activity accompanies neural circuit maturation throughout the developing brain1,2. The recent discovery of similar activity in the developing Drosophila central nervous system suggests that developmental activity is fundamental to the assembly of complex brains3. How such activity is coordinated across disparate brain regions to influence synaptic development at the level of defined cell types is not well understood. Here we show that neurons expressing the cation channel transient receptor potential gamma (Trpγ) relay and pattern developmental activity throughout the Drosophila brain. In trpγ mutants, activity is attenuated globally, and both patterns of activity and synapse structure are altered in a cell-type-specific manner. Less than 2% of the neurons in the brain express Trpγ. These neurons arborize throughout the brain, and silencing or activating them leads to loss or gain of brain-wide activity. Together, these results indicate that this small population of neurons coordinates brain-wide developmental activity. We propose that stereotyped patterns of developmental activity are driven by a discrete, genetically specified network to instruct neural circuit assembly at the level of individual cells and synapses. This work establishes the fly brain as an experimentally tractable system for studying how activity contributes to synapse and circuit formation