The Eph receptors are the largest known family of receptor tyrosine kinases. Initially all of them were identified as orphan receptors without known ligands, and their specific functions were not well understood. During the past few years, a corresponding family of ligands has been identified, called the ephrins, and specific functions have now been identified in neural development. The ephrins and Eph receptors are implicated as positional labels that may guide the development of neural topographic maps. They have also been implicated in pathway selection by axons, the guidance of cell migration, and the establishment of regional pattern in the nervous system. The ligands are anchored to cell surfaces, and most of the functions so far identified can be interpreted as precise guidance of cell or axon movement. This large family of ligands and receptors may make a major contribution to the accurate spatial patterning of connections and cell position in the nervous system.

Topographic maps with a defined spatial ordering of neuronal connections are a key feature of brain organization. Such maps are believed to develop in response to complementary position-specific labels in presynaptic and postsynaptic fields. However, the complementary labeling molecules are not known. In the well-studied visual map of retinal axons projecting to the tectum, the labels are hypothesized to be in gradients, without needing large numbers of cell-specific molecules. We recently cloned ELF-1 as a ligand for Eph family receptors. Here, RNA hybridization shows matching expression gradients for ELF-1 in the tectum and its receptor Mek4 in the retina. Binding activity detected with alkaline phosphatase fusions of ELF-1 and Mek4 also reveals gradients and provides direct evidence for molecular complementarity of gradients in reciprocal fields. ELF-1 and Mek4 may therefore play roles in retinotectal development and have properties predicted of topographic mapping labels.

Chondroitin sulfate proteoglycans (CSPGs) present a barrier to axon regeneration. However, no specific receptor for the inhibitory effect of CSPGs has been identified. We showed that a transmembrane protein tyrosine phosphatase, PTPσ, binds with high affinity to neural CSPGs. Binding involves the chondroitin sulfate chains and a specific site on the first immunoglobulin-like domain of PTPσ. In culture, PTPσ −/− neurons show reduced inhibition by CSPG. A PTPσ fusion protein probe can detect cognate ligands that are up-regulated specifically at neural lesion sites. After spinal cord injury, PTPσ gene disruption enhanced the ability of axons to penetrate regions containing CSPG. These results indicate that PTPσ can act as a receptor for CSPGs and may provide new therapeutic approaches to neural regeneration.Recovery after central nervous system (CNS) injury is minimal, leading to substantial current interest in potential strategies to overcome this challenge (1-5). Chondroitin sulfate proteoglycans (CSPGs) show dramatic up-regulation after neural injury, within the extracellular matrix of scar tissue and in the perineuronal net within more-distant targets of the severed axons (6,7). The inhibitory nature of CSPGs is reflected not only in the formation of dystrophic axonal retraction bulbs that fail to regenerate through the lesion (8), but also in the limited capacity for collateral sprouting of spared fibers (8,9). This inhibition can be relieved by chondroitinase ABC digestion of the chondroitin sulfate (CS) side chains, which can promote regeneration and sprouting and restore lost function (10)(11)(12)(13)(14). It has been known for nearly two decades that sulfated proteoglycans are major contributors to the repulsive nature of the glial scar (15); however, the precise inhibitory mechanism remains poorly understood. Because the identification of specific neuronal receptors for CSPGs has been lacking, relatively nonspecific mechanisms brought about by arrays of negatively charged sulfate (16) or the occlusion of substrate adhesion molecules (17) have been suggested.Transmembrane protein tyrosine phosphatases (PTPs) form a large and diverse molecular family and have a structure typical of transmembrane cell-surface receptors (18,19). In previous work, we and others have found that PTPσ and other PTPs in the leukocyte antigen-related (LAR) subfamily can act as receptors for heparan sulfate proteoglycans (HSPGs) (20)(21)(22), and these PTPs are involved in axon guidance and synapse formation during development (18- ‡To whom correspondence should be addressed. flanagan@hms.harvard.edu. * These authors contributed equally to this work. † Present address: Motor Neuron Center, Columbia University, New York, NY 10032, USA. (Fig. 1A). Using a cell-free system with recombinant fusion proteins of the PTPσ extra-cellular domain with an immunoglobulin Fc tag (PTPσ-Fc) and neurocan with an alkaline phosphatase tag (Ncn-AP), a binding interaction was indeed identified (P < 0.001) (Fig. 1B). Genuine biological ligand-re...

Ephrin-A2 and -A5 are thought to be anteroposterior mapping labels for the retinotectal/retinocollicular projection. Here, gene disruptions of both these ephrins are characterized. Focal retinal labeling reveals moderate map abnormalities when either gene is disrupted. Double heterozygotes also have a phenotype, showing an influence of absolute levels. In vitro assays indicate these ephrins are required for repellent activity in the target and also normal responsiveness in the retina. In double homozygotes, anteroposterior order is almost though not completely lost. Temporal or nasal retinal labelings reveal quantitatively similar but opposite shifts, with multiple terminations scattered widely over the target. These results indicate an axon competition mechanism for mapping, with a critical role for ephrins as anteroposterior topographic labels. Dorsoventral topography is also impaired, showing these ephrins are required in mapping both axes.

Heparin is required for the binding of basic fibroblast growth factor (bFGF) to high-affinity receptors on cells deficient in cell surface heparan sulfate proteoglycan. So that this heparin requirement could be evaluated in the absence of other cell surface molecules, we designed a simple assay based on a genetically engineered soluble form of murine FGF receptor 1 (mFRl) tagged with placental alkaline phosphatase. Using this assay, we showed that FGF-receptor binding has an absolute requirement for heparin. By using a cytokine-dependent lymphoid cell line engineered to express mFRl, we also showed that FGF-induced mitogenic activity is heparin dependent. Furthermore, we tested a series of small heparin oligosaccharides of defined lengths for their abilities to support bFGF-receptor binding and biologic activity. We found that a heparin oligosaccharide with as few as eight sugar residues is sufficient to support these activities. We also demonstrated that heparin facilitates FGF dimerization, a property that may be important for receptor activation.Heparin or heparan sulfate is required for basic fibroblast growth factor (bFGF) high-affinity receptor binding (38) and for bFGF-induced fibroblast growth and myoblast differentiation (29). These observations suggest that heparan sulfate proteoglycans (HSPGs) may be important regulators of bFGF biologic activity, acting directly at the level of the cell surface receptor (32). Not surprisingly, heparin and HSPGs are important regulators of cell growth (37). Depending on the tissue or cell type, heparin can either stimulate or inhibit cell proliferation (reviewed in references 14 and 31). Some of these effects may be mediated by FGF; however, other growth factors (including granulocyte-macrophage colonystimulating factor, [GM-CSF], interleukin 3 [IL-3], pleiotrophin, platelet factor 4, keratinocyte autocrine factor or amphiregulin, and heparin-binding-epidermal growth factor [EGF]) are also known to interact with heparin and may mediate some of these effects (6, 16; reviewed in reference 32). With the exception of FGF and amphiregulin, it is not known that heparin modulates growth factor activity at the level of a cell surface receptor. Nevertheless, heparin decreases the binding of amphiregulin to its receptor (6) and is required for bFGF-receptor binding (38). In addition, bFGF and the hematopoietic growth factors GM-CSF and IL-3, are biologically active when bound to HSPG (26,30,36).bFGF is known to reside in the extracellular matrix (ECM) of a wide variety of cells and tissues. This reservoir for bFGF serves to limit the diffusibility of the growth factor and thus to regulate its bioavailability. Biologically active bFGF can be displaced from the ECM by heparin or be released from the ECM by heparin-degrading enzymes (reviewed in references 14 and 20). Recently, syndecan, an HSPG, has been identified as a low-affinity binding site for bFGF (18 The observation that HSPGs serve as low-affinity binding sites for bFGF has been confirmed by comparing low-affinity FGF bi...

The model predicts that ONH biomechanics are strongly dependent on scleral biomechanical properties. Acute deformations of ONH tissues, and the consequent high levels of neural tissue strain, were less strongly dependent on the action of IOP directly on the internal surface of the ONH than on the indirect effects of IOP on the sclera. This suggests that interindividual variations in scleral properties could be a risk factor for the development of glaucoma. Eye size and lamina cribrosa biomechanical properties also have a strong influence on ONH biomechanics.

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Contact-mediated axon repulsion by ephrins raises an unresolved question: these cell surface ligands form a high-affinity multivalent complex with their receptors present on axons, yet rather than being bound, axons can be rapidly repelled. We show here that ephrin-A2 forms a stable complex with the metalloprotease Kuzbanian, involving interactions outside the cleavage region and the protease domain. Eph receptor binding triggered ephrin-A2 cleavage in a localized reaction specific to the cognate ligand. A cleavage-inhibiting mutation in ephrin-A2 delayed axon withdrawal. These studies reveal mechanisms for protease recognition and control of cell surface proteins, and, for ephrin-A2, they may provide a means for efficient axon detachment and termination of signaling.