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The Netrin receptor Dcc and its Drosophila homolog Frazzled play crucial roles in diverse developmental process, including axon guidance. In Drosophila, Fra regulates midline axon guidance through a Netrin-dependent and a Netrin-independent pathway. However, what molecules regulate these distinct signaling pathways remain unclear. To identify Fra-interacting proteins, we performed affinity purification mass spectrometry to establish a neuronalspecific Fra interactome. In addition to known interactors of Fra and Dcc, including Netrin and Robo1, our screen identified 85 candidate proteins, the majority of which are conserved in humans. Many of these proteins are expressed in the ventral nerve cord, and gene ontology, pathway analysis and biochemical validation identified several previously unreported pathways, including the receptor tyrosine phosphatase Lar, subunits of the COP9 signalosome and Rho-5, a regulator of the metalloprotease Tace. Finally, genetic analysis demonstrates that these genes regulate axon guidance and may define as yet unknown signaling mechanisms for Fra and its vertebrate homolog Dcc. Thus, the Fra interactome represents a resource to guide future functional studies. [ABSTRACT FROM AUTHOR]

Traumatic spinal cord injury produces long-term neurological damage, and presents a significant public health problem with nearly 18,000 new cases per year in the U.S. The injury results in both acute and chronic changes in the spinal cord, ultimately resulting in the production of a glial scar, consisting of multiple cells including fibroblasts, macrophages, microglia, and reactive astrocytes. Within the scar, there is an accumulation of extracellular matrix (ECM) molecules—primarily tenascins and chondroitin sulfate proteoglycans (CSPGs)—which are considered to be inhibitory to axonal regeneration. In this review article, we discuss the role of CSPGs in the injury response, especially how sulfated glycosaminoglycan (GAG) chains act to inhibit plasticity and regeneration. This includes how sulfation of GAG chains influences their biological activity and interactions with potential receptors. Comprehending the role of CSPGs in the inhibitory properties of the glial scar provides critical knowledge in the much-needed production of new therapies.

Traumatic spinal cord injury produces long-term neurological damage, and presents a significant public health problem with nearly 18,000 new cases per year in the U.S. The injury results in both acute and chronic changes in the spinal cord, ultimately resulting in the production of a glial scar, consisting of multiple cells including fibroblasts, macrophages, microglia, and reactive astrocytes. Within the scar, there is an accumulation of extracellular matrix (ECM) molecules—primarily tenascins and chondroitin sulfate proteoglycans (CSPGs)—which are considered to be inhibitory to axonal regeneration. In this review article, we discuss the role of CSPGs in the injury response, especially how sulfated glycosaminoglycan (GAG) chains act to inhibit plasticity and regeneration. This includes how sulfation of GAG chains influences their biological activity and interactions with potential receptors. Comprehending the role of CSPGs in the inhibitory properties of the glial scar provides critical knowledge in the much-needed production of new therapies. [ABSTRACT FROM AUTHOR]

Alternative splicing regulates trans‐synaptic adhesions and synapse development, but supporting in vivo evidence is limited. PTPδ, a receptor tyrosine phosphatase adhering to multiple synaptic adhesion molecules, is associated with various neuropsychiatric disorders; however, its in vivo functions remain unclear. Here, we show that PTPδ is mainly present at excitatory presynaptic sites by endogenous PTPδ tagging. Global PTPδ deletion in mice leads to input‐specific decreases in excitatory synapse development and strength. This involves tyrosine dephosphorylation and synaptic loss of IL1RAPL1, a postsynaptic partner of PTPδ requiring the PTPδ‐meA splice insert for binding. Importantly, PTPδ‐mutant mice lacking the PTPδ‐meA insert, and thus lacking the PTPδ interaction with IL1RAPL1 but not other postsynaptic partners, recapitulate biochemical and synaptic phenotypes of global PTPδ‐mutant mice. Behaviorally, both global and meA‐specific PTPδ‐mutant mice display abnormal sleep behavior and non‐REM rhythms. Therefore, alternative splicing in PTPδ regulates excitatory synapse development and sleep by modulating a specific trans‐synaptic adhesion. Synopsis: Global deletion of PTPδ, a presynaptic adhesion molecule, disrupts excitatory synapse formation and NREM sleep. Deletion of the PTPδ‐meA splice insert, critical for trans‐synaptic IL1RAPL1 binding, recapitulates the synaptic and behavioural changes induced by global PTPδ knockout. Global PTPδ deletion in mice leads to input‐specific decreases in excitatory synapse development and strength.This involves tyrosine dephosphorylation and synaptic loss of IL1RAPL1, a postsynaptic adhesion molecule binding to PTPδ with the meA splice insert.Deletion of the PTPδ‐meA splice insert recapitulates biochemical, synaptic, and behaviour changes induced by global PTPδ knockout. [ABSTRACT FROM AUTHOR]

The Netrin receptor Dcc and its Drosophila homolog Frazzled play crucial roles in diverse developmental process, including axon guidance. In Drosophila, Fra regulates midline axon guidance through a Netrin-dependent and a Netrin-independent pathway. However, what molecules regulate these distinct signaling pathways remain unclear. To identify Fra-interacting proteins, we performed affinity purification mass spectrometry to establish a neuronal-specific Fra interactome. In addition to known interactors of Fra and Dcc, including Netrin and Robo1, our screen identified 85 candidate proteins, the majority of which are conserved in humans. Many of these proteins are expressed in the ventral nerve cord, and gene ontology, pathway analysis and biochemical validation identified several previously unreported pathways, including the receptor tyrosine phosphatase Lar, subunits of the COP9 signalosome and Rho-5, a regulator of the metalloprotease Tace. Finally, genetic analysis demonstrates that these genes regulate axon guidance and may define as yet unknown signaling mechanisms for Fra and its vertebrate homolog Dcc. Thus, the Fra interactome represents a resource to guide future functional studies., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Sensory dendrite morphogenesis is directed by intrinsic and extrinsic factors. The extracellular environment plays instructive roles in patterning dendrite growth and branching. However, the molecular mechanism is not well understood. In Caenorhabditis elegans , the proprioceptive neuron PVD forms highly branched sensory dendrites adjacent to the hypodermis. We report that receptor tyrosine phosphatase CLR-1 functions in the hypodermis to pattern the PVD dendritic branches. Mutations in clr-1 lead to loss of quaternary branches, reduced secondary branches and increased ectopic branches. CLR-1 is necessary for the dendrite extension but not for the initial filopodia formation. Its role is dependent on the intracellular phosphatase domain but not the extracellular adhesion domain, indicating that it functions through dephosphorylating downstream factors but not through direct adhesion with neurons. Genetic analysis reveals that clr-1 also functions in parallel with SAX-7/DMA-1 pathway to control PVD primary dendrite development. We provide evidence of a new environmental factor for PVD dendrite morphogenesis. [ABSTRACT FROM AUTHOR]

Receptor-like protein tyrosine phosphatases represent a large protein family related to cell adhesion molecules, with diverse roles throughout neural development in vertebrates and invertebrates. This review focuses on their roles in axon growth, guidance and repair, as well as more recent findings demonstrating their key roles in pre-synaptic and post-synaptic maturation and function. These enzymes have been linked to memory and neuropsychiatric defects in loss-of-function rodent models, highlighting their potential as future drug targets. [ABSTRACT FROM AUTHOR]

Abstract: Receptor tyrosine phosphatases (RPTPs) are required for axon guidance during embryonic development in Drosophila. Here we examine the roles of four RPTPs during development of the larval mushroom body (MB). MB neurons extend axons into parallel tracts known as the peduncle and lobes. The temporal order of neuronal birth is reflected in the organization of axons within these tracts. Axons of the youngest neurons, known as core fibers, extend within a single bundle at the center, while those of older neurons fill the outer layers. RPTPs are selectively expressed on the core fibers of the MB. Ptp10D and Ptp69D regulate segregation of the young axons into a single core bundle. Ptp69D signaling is required for axonal extension beyond the peduncle. Lar and Ptp69D are necessary for the axonal branching decisions that create the lobes. Avoidance of the brain midline by extending medial lobe axons involves signaling through Lar. [Copyright &y& Elsevier]

Abstract: During gastrulation in Drosophila, ventral cells change shape, undergoing synchronous apical constriction, to create the ventral furrow (VF). This process is affected in mutant embryos lacking zygotic function of the folded gastrulation (fog) gene, which encodes a putative secreted protein. Fog is an essential autocrine signal that induces cytoskeletal changes in invaginating VF cells. Here we show that Fog is also required for nervous system development. Fog is expressed by longitudinal glia in the central nervous system (CNS), and reducing its expression in glia causes defects in process extension and axon ensheathment. Glial Fog overexpression produces a disorganized glial lattice. Fog has a distinct set of functions in CNS neurons. Our data show that reduction or overexpression of Fog in these neurons produces axon guidance phenotypes. Interestingly, these phenotypes closely resemble those seen in embryos with altered expression of the receptor tyrosine phosphatase PTP52F. We conducted epistasis experiments to define the genetic relationships between Fog and PTP52F, and the results suggest that PTP52F is a downstream component of the Fog signaling pathway in CNS neurons. We also found that Ptp52F mutants have early VF phenotypes like those seen in fog mutants. [Copyright &y& Elsevier]

In the Drosophila visual system, the color-sensing photoreceptors R7 and R8 project their axons to two distinct layers in the medulla. Loss of the receptor tyrosine phosphatase LAR from R7 photoreceptors causes their axons to terminate prematurely in the R8 layer. Here we identify a null mutation in the Liprin-α gene based on a similar R7 projection defect. Liprin-α physically interacts with the inactive D2 phosphatase domain of LAR, and this domain is also essential for R7 targeting. However, another LAR-dependent function, egg elongation, requires neither Liprin-α nor the LAR D2 domain. Although human and Caenorhabditis elegans Liprin-α proteins have been reported to control the localization of LAR, we find that LAR localizes to focal adhesions in Drosophila S2R+ cells and to photoreceptor growth cones in vivo independently of Liprin-α. In addition, Liprin-α overexpression or loss of function can affect R7 targeting in the complete absence of LAR. We conclude that Liprin-α does not simply act by regulating LAR localization but also has LAR-independent functions. [ABSTRACT FROM AUTHOR]

Abstract: The transmembrane forms of the protein tyrosine phosphatases (PTPs) are evolutionarily conserved and have been implicated in diverse cellular and developmental functions. Due to their structure, these enzymes are known as “receptor-like” PTPs (RPTPs), yet most remain as orphan receptors with no proven ligands yet. The question of whether or not such ligands exist is critical, because RPTP regulation by extracellular cues could be central to their signal transduction mechanisms. Identification of ligands has therefore been an important goal in the field. However, even with powerful genetic and molecular techniques at our disposal, this goal has frequently been met with frustration. This review focuses on methods used in the search for such ligands and primarily on a commonly employed technique named RAP in situ. RAP in situ has been used with some success in recent years and relies on the ability of isolated extracellular domains of receptors to still recognise their cognate ligand partners in vitro. For this technique, fusion proteins are made between RPTP ectodomains and placental alkaline phosphatase and these fusion proteins are used to probe tissue sections, live cultured cells or whole tissues. The binding sites of the proteins are visualised by alkaline phosphatase histochemistry, directly pinpointing the location of potential ligands. RAP in situ has helped to identify ligands for PTPζ/β and PTPσ, as well as locating several other putative ligand sites in tissues. We provide troubleshooting details for RAP in situ and describe some other complementary techniques that can be used. [Copyright &y& Elsevier]

Segmentation in vertebrate embryos is controlled by a biochemical oscillator ('segmentation clock') intrinsic to the cells in the unsegmented presomitic mesoderm, and is manifested in cyclic transcription of genes involved in establishing somite polarity and boundaries. We show that the receptor protein tyrosine phosphatase ψ (RPTPψ) gene is essential for normal functioning of the somitogenesis clock in zebrafish. We show that reduction of RPTPψ activity using morpholino antisense oligonucleotides results in severe disruption of the segmental pattern of the embryo, and loss of cyclic gene expression in the presomitic mesoderm. Analysis of cyclic genes in RPTPψ morphant embryos indicates an important requirement for RPTPψ in the control of the somitogenesis clock upstream of or in parallel with Delta/Notch signalling. Impairing RPTPψ activity also interferes with convergent extension during gastrulation. We discuss this dual requirement for RPTPψ in terms of potential functions in Notch and Wnt signalling. [ABSTRACT FROM AUTHOR]

To study the role of receptor tyrosine phosphatases in vertebrate development, in particular somitogenesis, we have cloned chick receptor tyrosine phosphatase Ψ (cRPTPΨ). cRPTPΨ is expressed in a dynamic fashion in the somites to-be-formed and uniformly throughout the presomitic mesoderm. In differentiating somites, cRPTPΨ expression gets restricted to the dermomyotome. In addition cRPTPΨ is expressed in the developing intermediate mesoderm, in neurogenic and sensory organs, the limb bud and the developing heart. [Copyright &y& Elsevier]

Receptor protein-tyrosine phosphatase RPTPσ has important functions in modulating neural development and regeneration. Compelling evidence suggests that both heparan sulfate (HS) and chondroitin sulfate (CS) glycosaminoglycans (GAGs) bind to a series of Lys residues located in the first Ig domain of RPTPσ. However, HS promotes and CS inhibits axonal growth. Mutation of these Lys residues abolished binding and signal transduction of RPTPσ to CS, whereas HS binding was reduced, and signaling persisted. This activity was mediated through novel heparin-binding sites identified in the juxtamembrane region. Although different functional outcomes of HS and CS have been previously attributed to the differential oligomeric state of RPTPσ upon GAG binding, we found that RPTPσ was clustered by both heparin and CS GAG rich in 4,6- O -disulfated disaccharide units. We propose an additional mechanism by which RPTPσ distinguishes between HS and CS through these novel binding sites.

Tyrosine phosphorylation is at the crossroads of many signaling pathways. Brain wiring is not an exception, and several receptor tyrosine kinases (RTKs) and tyrosine receptor phosphates (RPTPs) have been involved in this process. Considerable work has been done on RTKs, and for many of them, detailed molecular mechanisms and functions in several systems have been characterized. In contrast, RPTPs have been studied considerably less and little is known about their ligands and substrates. In both families, we find redundancy between different members to accomplish particular wiring patterns. Strikingly, some RTKs and RPTPs have lost their catalytic activity during evolution, but not their importance in biological processes. In this regard, we have to keep in mind that these proteins have multiple domains and some of their functions are independent of tyrosine phosphorylation/dephosphorylation. Since RTKs and RPTPs are enzymes involved not only in early stages of axon and dendrite pathfinding but also in synapse formation and physiology, they have a potential as drug targets. Drosophila has been a key model organism in the search of a better understanding of brain wiring, and its sophisticated toolbox is very suitable for studying the function of genes with pleiotropic functions such as RTKs and RPTPs, from wiring to synaptic formation and function. In these review, we mainly cover findings from this model organism and complement them with discoveries in vertebrate systems., (© 2017 Elsevier Inc. All rights reserved.)

Receptor protein tyrosine phosphatases (RPTPs) form a group of over 20 enzymes in vertebrates, each with unique ectodomains subject to potential extracellular interactions with ligands. It has recently become clear that a remarkably diverse range of ligands exist, including homophilic binders, adhesion molecules, neurotrophin receptors, and proteoglycans. Individual RPTPs can bind several ligands, and vice versa, suggesting that complex cell signaling networks exist. The identification of RPTP ligands and where they are located in tissues remains a challenge for a large number of these enzymes. Here we describe some powerful methods that have proved successful for several research groups, leading to our improved understanding of RPTP-ligand interactions and functional regulation.