Human photoreceptors switch from autonomous axon extension to cell-mediated process pulling during synaptic marker redistribution.

Photoreceptors (PRs) are the primary visual sensory cells, and their loss leads to blindness that is currently incurable. Although cell replacement therapy holds promise, success is hindered by our limited understanding of PR axon growth during development and regeneration. Here, we generate retinal organoids from human pluripotent stem cells to study the mechanisms of PR process extension.

Growth Factors as Axon Guidance Molecules: Lessons From Studies.

Growth cones at the tips of extending axons navigate through developing organisms by probing extracellular cues, which guide them through intermediate steps and onto final synaptic target sites. Widespread focus on a few guidance cue families has historically overshadowed potentially crucial roles of less well-studied growth factors in axon guidance. In fact, recent evidence suggests that a variety of growth factors have the ability to guide axons, affecting the targeting and morphogenesis of growth cones This review summarizes experiments identifying responses and signaling mechanisms underlying axon morphogenesis caused by underappreciated growth factors.

RHOA signaling defects result in impaired axon guidance in iPSC-derived neurons from patients with tuberous sclerosis complex.

Patients with Tuberous Sclerosis Complex (TSC) show aberrant wiring of neuronal connections formed during development which may contribute to symptoms of TSC, such as intellectual disabilities, autism, and epilepsy. Yet models examining the molecular basis for axonal guidance defects in developing human neurons have not been developed. Here, we generate human induced pluripotent stem cell (hiPSC) lines from a patient with TSC and genetically engineer counterparts and isogenic controls.

Familiar growth factors have diverse roles in neural network assembly.

The assembly of neuronal circuits during development depends on guidance of axonal growth cones by molecular cues deposited in their environment. While a number of families of axon guidance molecules have been identified and reviewed, important and diverse activities of traditional growth factors are emerging. Besides clear and well recognized roles in the regulation of cell division, differentiation and survival, new research shows later phase roles for a number of growth factors in promoting neuronal migration, axon guidance and synapse formation throughout the nervous system.

Glial Cell-Axonal Growth Cone Interactions in Neurodevelopment and Regeneration.

The developing nervous system is a complex yet organized system of neurons, glial support cells, and extracellular matrix that arranges into an elegant, highly structured network. The extracellular and intracellular events that guide axons to their target locations have been well characterized in many regions of the developing nervous system. However, despite extensive work, we have a poor understanding of how axonal growth cones interact with surrounding glial cells to regulate network assembly.

Environmental Elasticity Regulates Cell-type Specific RHOA Signaling and Neuritogenesis of Human Neurons.

The microenvironment of developing neurons is a dynamic landscape of both chemical and mechanical cues that regulate cell proliferation, differentiation, migration, and axon extension. While the regulatory roles of chemical ligands in neuronal morphogenesis have been described, little is known about how mechanical forces influence neurite development. Here, we tested how substratum elasticity regulates neurite development of human forebrain (hFB) neurons and human motor neurons (hMNs), two populations of neurons that naturally extend axons into distinct elastic environments.

Calpain-Mediated Proteolysis of Talin and FAK Regulates Adhesion Dynamics Necessary for Axon Guidance.

Guidance of axons to their proper synaptic target sites requires spatially and temporally precise modulation of biochemical signals within growth cones. Ionic calcium (Ca) is an essential signal for axon guidance that mediates opposing effects on growth cone motility. The diverse effects of Ca arise from the precise localization of Ca signals into microdomains containing specific Ca effectors.

Division of labor in the growth cone by DSCR1.

Local protein synthesis directs growth cone turning of nascent axons, but mechanisms governing this process within compact, largely autonomous microenvironments remain poorly understood. In this issue, Wang et al. (2016.

Cell adhesion and invasion mechanisms that guide developing axons.

Axon extension, guidance and tissue invasion share many similarities to normal cell migration and cancer cell metastasis. Proper cell and growth cone migration requires tightly regulated adhesion complex assembly and detachment from the extracellular matrix (ECM). In addition, many cell types actively remodel the ECM using matrix metalloproteases (MMPs) to control tissue invasion and cell dispersal.

Guidance of Axons by Local Coupling of Retrograde Flow to Point Contact Adhesions.

Unlabelled: Growth cones interact with the extracellular matrix (ECM) through integrin receptors at adhesion sites termed point contacts. Point contact adhesions link ECM proteins to the actin cytoskeleton through numerous adaptor and signaling proteins. One presumed function of growth cone point contacts is to restrain or "clutch" myosin-II-based filamentous actin (F-actin) retrograde flow (RF) to promote leading edge membrane protrusion.

Mechanochemical regulation of growth cone motility.

Neuronal growth cones are exquisite sensory-motor machines capable of transducing features contacted in their local extracellular environment into guided process extension during development. Extensive research has shown that chemical ligands activate cell surface receptors on growth cones leading to intracellular signals that direct cytoskeletal changes. However, the environment also provides mechanical support for growth cone adhesion and traction forces that stabilize leading edge protrusions.

Regulation of ECM degradation and axon guidance by growth cone invadosomes.

Invadopodia and podosomes, collectively referred to as invadosomes, are F-actin-rich basal protrusions of cells that provide sites of attachment to and degradation of the extracellular matrix. Invadosomes promote the invasion of cells, ranging from metastatic cancer cells to immune cells, into tissue. Here, we show that neuronal growth cones form protrusions that share molecular, structural and functional characteristics of invadosomes.

iPSC-derived forebrain neurons from FXS individuals show defects in initial neurite outgrowth.

Fragile X syndrome (FXS) is the most common form of inherited intellectual disability and is closely linked with autism. The genetic basis of FXS is an expansion of CGG repeats in the 5'-untranslated region of the FMR1 gene on the X chromosome leading to the loss of expression of the fragile X mental retardation protein (FMRP). The cause of FXS has been known for over 20 years, yet the full molecular and cellular consequences of this mutation remain unclear.

Actin dynamics in growth cone motility and navigation.

Motile growth cones lead growing axons through developing tissues to synaptic targets. These behaviors depend on the organization and dynamics of actin filaments that fill the growth cone leading margin [peripheral (P-) domain]. Actin filament organization in growth cones is regulated by actin-binding proteins that control all aspects of filament assembly, turnover, interactions with other filaments and cytoplasmic components, and participation in producing mechanical forces.

CIP4 coordinates with phospholipids and actin-associated proteins to localize to the protruding edge and produce actin ribs and veils.

Cdc42-interacting protein 4 (CIP4), a member of the F-BAR family of proteins, plays important roles in a variety of cellular events by regulating both membrane and actin dynamics. In many cell types, CIP4 functions in vesicle formation, endocytosis and membrane tubulation. However, recent data indicate that CIP4 is also involved in protrusion in some cell types, including cancer cells (lamellipodia and invadopodia) and neurons (ribbed lamellipodia and veils).

PAK-PIX interactions regulate adhesion dynamics and membrane protrusion to control neurite outgrowth.

The roles of P21-activated kinase (PAK) in the regulation of axon outgrowth downstream of extracellular matrix (ECM) proteins are poorly understood. Here we show that PAK1-3 and PIX are expressed in the developing spinal cord and differentially localize to point contacts and filopodial tips within motile growth cones. Using a specific interfering peptide called PAK18, we found that axon outgrowth is robustly stimulated on laminin by partial inhibition of PAK-PIX interactions and PAK function, whereas complete inhibition of PAK function stalls axon outgrowth.

Mechanosensitive TRPC1 channels promote calpain proteolysis of talin to regulate spinal axon outgrowth.

Intracellular Ca(2+) signals control the development and regeneration of spinal axons downstream of chemical guidance cues, but little is known about the roles of mechanical cues in axon guidance. Here we show that transient receptor potential canonical 1 (TRPC1) subunits assemble mechanosensitive (MS) channels on Xenopus neuronal growth cones that regulate the extension and direction of axon outgrowth on rigid, but not compliant, substrata. Reducing expression of TRPC1 by antisense morpholinos inhibits the effects of MS channel blockers on axon outgrowth and local Ca(2+) transients.

Focal adhesion kinase modulates Cdc42 activity downstream of positive and negative axon guidance cues.

There is biochemical, imaging and functional evidence that Rho GTPase signaling is a crucial regulator of actin-based structures such as lamellipodia and filopodia. However, although Rho GTPases are believed to serve similar functions in growth cones, the spatiotemporal dynamics of Rho GTPase signaling has not been examined in living growth cones in response to known axon guidance cues. Here we provide the first measurements of Cdc42 activity in living growth cones acutely stimulated with both growth-promoting and growth-inhibiting axon-guidance cues.

Focal adhesion kinase promotes integrin adhesion dynamics necessary for chemotropic turning of nerve growth cones.

The ability of extending axons to navigate using combinations of extracellular cues is essential for proper neural network formation. One intracellular signaling molecule that integrates convergent signals from both extracellular matrix (ECM) proteins and growth factors is focal adhesion kinase (FAK). Analysis of FAK function shows that it influences a variety of cellular activities, including cell motility, proliferation, and differentiation.

Pioneering studies on the mechanisms of neuronal morphogenesis.

Axon outgrowth and pathfinding occurs through a complex series of interacting biochemical signaling pathways that regulate the motility of neuronal growth cones. Over the past 30 years, Paul Letourneau and his students have explored the molecular basis of growth cone motility and have contributed immensely to this field. In celebration of his 65th birthday, this essay is written in gratitude for Paul's many contributions and training.

Regulation of axonal outgrowth and pathfinding by integrin-ECM interactions.

Developing neurons use a combination of guidance cues to assemble a functional neural network. A variety of proteins immobilized within the extracellular matrix (ECM) provide specific binding sites for integrin receptors on neurons. Integrin receptors on growth cones associate with a number of cytosolic adaptor and signaling proteins that regulate cytoskeletal dynamics and cell adhesion.

Imaging adhesion and signaling dynamics in Xenopus laevis growth cones.

Xenopus laevis provides a robust model system to study cellular signaling and downstream processes during development both in vitro and in vivo. Intracellular signals must function within highly restricted spatial and temporal domains to activate specific downstream targets and cellular processes. Combining the versatility of developing Xenopus neurons with advances in fluorescent protein biosensors and imaging technologies has allowed many dynamic cellular processes to be visualized.

Balanced Vav2 GEF activity regulates neurite outgrowth and branching in vitro and in vivo.

We have investigated the role of Vav2, a reported Rac1/Cdc42 GEF, on the development of Xenopus spinal neurons in vitro and in vivo. Both gain and loss of Vav2 function inhibited the rate neurite extension on laminin (LN), while only GFP-Vav2 over-expression enhanced process formation and branching. Vav2 over-expression protected neurons from RhoA-mediated growth cone collapse, similar to constitutively active Rac1, suggesting that Vav2 activates Rac1 in spinal neurons.

Retinotopic mapping requires focal adhesion kinase-mediated regulation of growth cone adhesion.

Adhesion controls growth cone motility, yet the effects of axon guidance cues on adhesion site dynamics are poorly understood. Here we show that ephrin-A1 reduces retinal ganglion cell (RGC) axon outgrowth by stabilizing existing adhesions and inhibiting new adhesion assembly. Ephrin-A1 activates focal adhesion kinase (FAK) in an integrin- and Src-dependent manner and the effects of ephrin-A1 on growth cone motility require FAK activation.