Slit and Receptor Tyrosine Phosphatase 69D Confer Spatial Specificity to Axon Branching via Dscam1.

Axonal branching contributes substantially to neuronal circuit complexity. Studies in Drosophila have shown that loss of Dscam1 receptor diversity can fully block axon branching in mechanosensory neurons. Here we report that cell-autonomous loss of the receptor tyrosine phosphatase 69D (RPTP69D) and loss of midline-localized Slit inhibit formation of specific axon collaterals through modulation of Dscam1 activity. Genetic and biochemical data support a model in which direct binding of Slit to Dscam1 enhances the interaction of Dscam1 with RPTP69D, stimulating Dscam1 dephosphorylation. Single-growth-cone imaging reveals that Slit/RPTP69D are not required for general branch initiation but instead promote the extension of specific axon collaterals. Hence, although regulation of intrinsic Dscam1-Dscam1 isoform interactions is essential for formation of all mechanosensory-axon branches, the local ligand-induced alterations of Dscam1 phosphorylation in distinct growth-cone compartments enable the spatial specificity of axon collateral formation.

Fat2 polarizes Lar and Sema5c to coordinate the motility of collectively migrating epithelial cells.

Migrating epithelial cells globally align their migration machinery to achieve tissue-level movement. Biochemical signaling across leading-trailing cell-cell interfaces can promote this alignment by partitioning migratory behaviors like protrusion and retraction to opposite sides of the interface. However, how signaling proteins become organized at interfaces to accomplish this is poorly understood. The follicular epithelial cells of Drosophila melanogaster have two signaling modules at their leading-trailing interfaces - one composed of the atypical cadherin Fat2 (also known as Kugelei) and the receptor tyrosine phosphatase Lar, and one composed of Semaphorin5c and its receptor Plexin A. Here, we show that these modules form one interface signaling system with Fat2 at its core. Trailing edge-enriched Fat2 concentrates both Lar and Semaphorin5c at leading edges of cells, but Lar and Semaphorin5c play little role in the localization of Fat2. Fat2 is also more stable at interfaces than Lar or Semaphorin5c. Once localized, Lar and Semaphorin5c act in parallel to promote collective migration. We propose that Fat2 serves as the organizer of this interface signaling system by coupling and polarizing the distributions of multiple effectors that work together to align the migration machinery of neighboring cells.

Systematic analysis of the Frazzled receptor interactome establishes previously unreported regulators of axon guidance.

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.

Protein tyrosine phosphatase 69D is a substrate of protein O-mannosyltransferases 1-2 that is required for the wiring of sensory axons in Drosophila.

Mutations in protein O-mannosyltransferases (POMTs) result in severe brain defects and congenital muscular dystrophies characterized by abnormal glycosylation of α-dystroglycan (α-Dg). However, neurological phenotypes of POMT mutants are not well understood, and the functional substrates of POMTs other than α-Dg remain unknown. Using a Drosophila model, here we reveal that Dg alone cannot account for the phenotypes of POMT mutants, and identify Protein tyrosine phosphatase 69D (PTP69D) as a gene interacting with POMTs in producing the abdomen rotation phenotype. Using RNAi-mediated knockdown, mutant alleles, and a dominant-negative form of PTP69D, we reveal that PTP69D is required for the wiring of larval sensory axons. We also found that PTP69D and POMT genes interact in this process, and that their interactions lead to complex synergistic or antagonistic effects on axon wiring phenotypes, depending on the mode of genetic manipulation. Using glycoproteomic approaches, we further characterized the glycosylation of the PTP69D transgenic construct expressed in genetic strains with different levels of POMT activity. We found that the PTP69D construct carries many O-linked mannose modifications when expressed in Drosophila with wild-type or ectopically upregulated expression of POMTs. These modifications were absent in POMT mutants, suggesting that PTP69D is a substrate of POMT-mediated O-mannosylation. Taken together, our results indicate that PTP69D is a novel functional substrate of POMTs that is required for axon connectivity. This mechanism of POMT-mediated regulation of receptor-type protein tyrosine phosphatase functions could potentially be conserved in mammals and may shed new light on the etiology of neurological defects in muscular dystrophies.

Lar maintains the homeostasis of the hematopoietic organ in Drosophila by regulating insulin signaling in the niche.

Stem cell compartments in metazoa get regulated by systemic factors as well as local stem cell niche-derived factors. However, the mechanisms by which systemic signals integrate with local factors in maintaining tissue homeostasis remain unclear. Employing the Drosophila lymph gland, which harbors differentiated blood cells, and stem-like progenitor cells and their niche, we demonstrate how a systemic signal interacts and harmonizes with local factor/s to achieve cell type-specific tissue homeostasis. Our genetic analyses uncovered a novel function of Lar, a receptor protein tyrosine phosphatase. Niche-specific loss of Lar leads to upregulated insulin signaling, causing increased niche cell proliferation and ectopic progenitor differentiation. Insulin signaling assayed by PI3K activation is downregulated after the second instar larval stage, a time point that coincides with the appearance of Lar in the hematopoietic niche. We further demonstrate that Lar physically associates with InR and serves as a negative regulator for insulin signaling in the Drosophila larval hematopoietic niche. Whether Lar serves as a localized invariable negative regulator of systemic signals such as insulin in other stem cell niches remains to be explored.

LAR-RPTPs Directly Interact with Neurexins to Coordinate Bidirectional Assembly of Molecular Machineries.

Neurexins (Nrxns) and LAR-RPTPs (leukocyte common antigen-related protein tyrosine phosphatases) are presynaptic adhesion proteins responsible for organizing presynaptic machineries through interactions with nonoverlapping extracellular ligands. Here, we report that two members of the LAR-RPTP family, PTPσ and PTPδ, are required for the presynaptogenic activity of Nrxns. Intriguingly, Nrxn1 and PTPσ require distinct sets of intracellular proteins for the assembly of specific presynaptic terminals. In addition, Nrxn1α showed robust heparan sulfate (HS)-dependent, high-affinity interactions with Ig domains of PTPσ that were regulated by the splicing status of PTPσ. Furthermore, Nrxn1α WT, but not a Nrxn1α mutant lacking HS moieties (Nrxn1α ΔHS), inhibited postsynapse-inducing activity of PTPσ at excitatory, but not inhibitory, synapses. Similarly, cis expression of Nrxn1α WT, but not Nrxn1α ΔHS, suppressed the PTPσ-mediated maintenance of excitatory postsynaptic specializations in mouse cultured hippocampal neurons. Lastly, genetics analyses using male or female Drosophila Dlar and Dnrx mutant larvae identified epistatic interactions that control synapse formation and synaptic transmission at neuromuscular junctions. Our results suggest a novel synaptogenesis model whereby different presynaptic adhesion molecules combine with distinct regulatory codes to orchestrate specific synaptic adhesion pathways.SIGNIFICANCE STATEMENT We provide evidence supporting the physical interactions of neurexins with leukocyte common-antigen related receptor tyrosine phosphatases (LAR-RPTPs). The availability of heparan sulfates and alternative splicing of LAR-RPTPs regulate the binding affinity of these interactions. A set of intracellular presynaptic proteins is involved in common for Nrxn- and LAR-RPTP-mediated presynaptic assembly. PTPσ triggers glutamatergic and GABAergic postsynaptic differentiation in an alternative splicing-dependent manner, whereas Nrxn1α induces GABAergic postsynaptic differentiation in an alternative splicing-independent manner. Strikingly, Nrxn1α inhibits the glutamatergic postsynapse-inducing activity of PTPσ, suggesting that PTPσ and Nrxn1α might control recruitment of a different pool of postsynaptic machinery. Drosophila orthologs of Nrxns and LAR-RPTPs mediate epistatic interactions in controlling synapse structure and strength at neuromuscular junctions, underscoring the physiological significance in vivo.

The receptor protein tyrosine phosphatase CLR-1 is required for synaptic partner recognition.

During neural circuit formation, most axons are guided to complex environments, coming into contact with multiple potential synaptic partners. However, it is critical that they recognize specific neurons with which to form synapses. Here, we utilize the split GFP-based marker Neuroligin-1 GFP Reconstitution Across Synaptic Partners (NLG-1 GRASP) to visualize specific synapses in live animals, and a circuit-specific behavioral assay to probe circuit function. We demonstrate that the receptor protein tyrosine phosphatase (RPTP) clr-1 is necessary for synaptic partner recognition (SPR) between the PHB sensory neurons and the AVA interneurons in C. elegans. Mutations in clr-1/RPTP result in reduced NLG-1 GRASP fluorescence and impaired behavioral output of the PHB circuit. Temperature-shift experiments demonstrate that clr-1/RPTP acts early in development, consistent with a role in SPR. Expression and cell-specific rescue experiments indicate that clr-1/RPTP functions in postsynaptic AVA neurons, and overexpression of clr-1/RPTP in AVA neurons is sufficient to direct additional PHB-AVA synaptogenesis. Genetic analysis reveals that clr-1/RPTP acts in the same pathway as the unc-6/Netrin ligand and the unc-40/DCC receptor, which act in AVA and PHB neurons, respectively. This study defines a new mechanism by which SPR is governed, and demonstrates that these three conserved families of molecules, with roles in neurological disorders and cancer, can act together to regulate communication between cells.

Type IIa RPTPs and Glycans: Roles in Axon Regeneration and Synaptogenesis.

Type IIa receptor tyrosine phosphatases (RPTPs) play pivotal roles in neuronal network formation. It is emerging that the interactions of RPTPs with glycans, i.e., chondroitin sulfate (CS) and heparan sulfate (HS), are critical for their functions. We highlight here the significance of these interactions in axon regeneration and synaptogenesis. For example, PTPσ, a member of type IIa RPTPs, on axon terminals is monomerized and activated by the extracellular CS deposited in neural injuries, dephosphorylates cortactin, disrupts autophagy flux, and consequently inhibits axon regeneration. In contrast, HS induces PTPσ oligomerization, suppresses PTPσ phosphatase activity, and promotes axon regeneration. PTPσ also serves as an organizer of excitatory synapses. PTPσ and neurexin bind one another on presynapses and further bind to postsynaptic leucine-rich repeat transmembrane protein 4 (LRRTM4). Neurexin is now known as a heparan sulfate proteoglycan (HSPG), and its HS is essential for the binding between these three molecules. Another HSPG, glypican 4, binds to presynaptic PTPσ and postsynaptic LRRTM4 in an HS-dependent manner. Type IIa RPTPs are also involved in the formation of excitatory and inhibitory synapses by heterophilic binding to a variety of postsynaptic partners. We also discuss the important issue of possible mechanisms coordinating axon extension and synapse formation.

Understanding the causes of errors in eukaryotic protein-coding gene prediction: a case study of primate proteomes.

BACKGROUND: Recent advances in sequencing technologies have led to an explosion in the number of genomes available, but accurate genome annotation remains a major challenge. The prediction of protein-coding genes in eukaryotic genomes is especially problematic, due to their complex exon-intron structures. Even the best eukaryotic gene prediction algorithms can make serious errors that will significantly affect subsequent analyses. RESULTS: We first investigated the prevalence of gene prediction errors in a large set of 176,478 proteins from ten primate proteomes available in public databases. Using the well-studied human proteins as a reference, a total of 82,305 potential errors were detected, including 44,001 deletions, 27,289 insertions and 11,015 mismatched segments where part of the correct protein sequence is replaced with an alternative erroneous sequence. We then focused on the mismatched sequence errors that cause particular problems for downstream applications. A detailed characterization allowed us to identify the potential causes for the gene misprediction in approximately half (5446) of these cases. As a proof-of-concept, we also developed a simple method which allowed us to propose improved sequences for 603 primate proteins. CONCLUSIONS: Gene prediction errors in primate proteomes affect up to 50% of the sequences. Major causes of errors include undetermined genome regions, genome sequencing or assembly issues, and limitations in the models used to represent gene exon-intron structures. Nevertheless, existing genome sequences can still be exploited to improve protein sequence quality. Perspectives of the work include the characterization of other types of gene prediction errors, as well as the development of a more comprehensive algorithm for protein sequence error correction.

The secreted MSP domain of C. elegans VAPB homolog VPR-1 patterns the adult striated muscle mitochondrial reticulum via SMN-1.

The major sperm protein domain (MSPd) has an extracellular signaling function implicated in amyotrophic lateral sclerosis. Secreted MSPds derived from the C. elegans VAPB homolog VPR-1 promote mitochondrial localization to actin-rich I-bands in body wall muscle. Here we show that the nervous system and germ line are key MSPd secretion tissues. MSPd signals are transduced through the CLR-1 Lar-like tyrosine phosphatase receptor. We show that CLR-1 is expressed throughout the muscle plasma membrane, where it is accessible to MSPd within the pseudocoelomic fluid. MSPd signaling is sufficient to remodel the muscle mitochondrial reticulum during adulthood. An RNAi suppressor screen identified survival of motor neuron 1 (SMN-1) as a downstream effector. SMN-1 acts in muscle, where it colocalizes at myofilaments with ARX-2, a component of the Arp2/3 actin-nucleation complex. Genetic studies suggest that SMN-1 promotes Arp2/3 activity important for localizing mitochondria to I-bands. Our results support the model that VAPB homologs are circulating hormones that pattern the striated muscle mitochondrial reticulum. This function is crucial in adults and requires SMN-1 in muscle, likely independent of its role in pre-mRNA splicing.

Demystifying the extracellular matrix and its proteolytic remodeling in the brain: structural and functional insights.

The extracellular matrix (ECM) plays diverse roles in several physiological and pathological conditions. In the brain, the ECM is unique both in its composition and in functions. Furthermore, almost all the cells in the central nervous system contribute to different aspects of this intricate structure. Brain ECM, enriched with proteoglycans and other small proteins, aggregate into distinct structures around neurons and oligodendrocytes. These special structures have cardinal functions in the normal functioning of the brain, such as learning, memory, and synapse regulation. In this review, we have compiled the current knowledge about the structure and function of important ECM molecules in the brain and their proteolytic remodeling by matrix metalloproteinases and other enzymes, highlighting the special structures they form. In particular, the proteoglycans in brain ECM, which are essential for several vital functions, are emphasized in detail.

Dendritic space-filling requires a neuronal type-specific extracellular permissive signal in Drosophila.

Neurons sometimes completely fill available space in their receptive fields with evenly spaced dendrites to uniformly sample sensory or synaptic information. The mechanisms that enable neurons to sense and innervate all space in their target tissues are poorly understood. Using Drosophila somatosensory neurons as a model, we show that heparan sulfate proteoglycans (HSPGs) Dally and Syndecan on the surface of epidermal cells act as local permissive signals for the dendritic growth and maintenance of space-filling nociceptive C4da neurons, allowing them to innervate the entire skin. Using long-term time-lapse imaging with intact Drosophila larvae, we found that dendrites grow into HSPG-deficient areas but fail to stay there. HSPGs are necessary to stabilize microtubules in newly formed high-order dendrites. In contrast to C4da neurons, non-space-filling sensory neurons that develop in the same microenvironment do not rely on HSPGs for their dendritic growth. Furthermore, HSPGs do not act by transporting extracellular diffusible ligands or require leukocyte antigen-related (Lar), a receptor protein tyrosine phosphatase (RPTP) and the only known Drosophila HSPG receptor, for promoting dendritic growth of space-filling neurons. Interestingly, another RPTP, Ptp69D, promotes dendritic growth of C4da neurons in parallel to HSPGs. Together, our data reveal an HSPG-dependent pathway that specifically allows dendrites of space-filling neurons to innervate all target tissues in Drosophila.

Receptor-like cytoplasmic kinase MARIS functions downstream of CrRLK1L-dependent signaling during tip growth.

Growing plant cells need to rigorously coordinate external signals with internal processes. For instance, the maintenance of cell wall (CW) integrity requires the coordination of CW sensing with CW remodeling and biosynthesis to avoid growth arrest or integrity loss. Despite the involvement of receptor-like kinases (RLKs) of the Catharanthus roseus RLK1-like (CrRLK1L) subfamily and the reactive oxygen species-producing NADPH oxidases, it remains largely unknown how this coordination is achieved. ANXUR1 (ANX1) and ANX2, two redundant members of the CrRLK1L subfamily, are required for tip growth of the pollen tube (PT), and their closest homolog, FERONIA, controls root-hair tip growth. Previously, we showed that ANX1 overexpression mildly inhibits PT growth by oversecretion of CW material, whereas pollen tubes of anx1 anx2 double mutants burst spontaneously after germination. Here, we report the identification of suppressor mutants with improved fertility caused by the rescue of anx1 anx2 pollen tube bursting. Mapping of one these mutants revealed an R240C nonsynonymous substitution in the activation loop of a receptor-like cytoplasmic kinase (RLCK), which we named MARIS (MRI). We show that MRI is a plasma membrane-localized member of the RLCK-VIII subfamily and is preferentially expressed in both PTs and root hairs. Interestingly, mri-knockout mutants display spontaneous PT and root-hair bursting. Moreover, expression of the MRI(R240C) mutant, but not its wild-type form, partially rescues the bursting phenotypes of anx1 anx2 PTs and fer root hairs but strongly inhibits wild-type tip growth. Thus, our findings identify a novel positive component of the CrRLK1L-dependent signaling cascade that coordinates CW integrity and tip growth.

Targeting Receptor-Type Protein Tyrosine Phosphatases with Biotherapeutics: Is Outside-in Better than Inside-Out?

Protein tyrosine phosphatases (PTPs), of the receptor and non-receptor classes, are key signaling molecules that play critical roles in cellular regulation underlying diverse physiological events. Aberrant signaling as a result of genetic mutation or altered expression levels has been associated with several diseases and treatment via pharmacological intervention at the level of PTPs has been widely explored; however, the challenges associated with development of small molecule phosphatase inhibitors targeting the intracellular phosphatase domain (the "inside-out" approach) have been well documented and as yet there are no clinically approved drugs targeting these enzymes. The alternative approach of targeting receptor PTPs with biotherapeutic agents (such as monoclonal antibodies or engineered fusion proteins; the "outside-in" approach) that interact with the extracellular ectodomain offers many advantages, and there have been a number of exciting recent developments in this field. Here we provide a brief overview of the receptor PTP family and an update on the emerging area of receptor PTP-targeted biotherapeutics for CD148, vascular endothelial-protein tyrosine phosphatase (VE-PTP), receptor-type PTPs σ, γ, ζ (RPTPσ, RPTPγ, RPTPζ) and CD45, and discussion of future potential in this area.

The Drosophila Receptor Protein Tyrosine Phosphatase LAR Is Required for Development of Circadian Pacemaker Neuron Processes That Support Rhythmic Activity in Constant Darkness But Not during Light/Dark Cycles.

InDrosophila, a transcriptional feedback loop that is activated by CLOCK-CYCLE (CLK-CYC) complexes and repressed by PERIOD-TIMELESS (PER-TIM) complexes keeps circadian time. The timing of CLK-CYC activation and PER-TIM repression is regulated post-translationally, in part through rhythmic phosphorylation of CLK, PER, and TIM. Although kinases that control PER, TIM, and CLK levels, activity, and/or subcellular localization have been identified, less is known about phosphatases that control clock protein dephosphorylation. To identify clock-relevant phosphatases, clock-cell-specific RNAi knockdowns ofDrosophilaphosphatases were screened for altered activity rhythms. One phosphatase that was identified, the receptor protein tyrosine phosphatase leukocyte-antigen-related (LAR), abolished activity rhythms in constant darkness (DD) without disrupting the timekeeping mechanism in brain pacemaker neurons. However, expression of the neuropeptide pigment-dispersing factor (PDF), which mediates pacemaker neuron synchrony and output, is eliminated in the dorsal projections from small ventral lateral (sLNv) pacemaker neurons whenLarexpression is knocked down during development, but not in adults. Loss ofLarfunction eliminates sLNvdorsal projections, but PDF expression persists in sLNvand large ventral lateral neuron cell bodies and their remaining projections. In contrast to the defects in lights-on and lights-off anticipatory activity seen in flies that lack PDF,LarRNAi knockdown flies anticipate the lights-on and lights-off transition normally. Our results demonstrate thatLaris required for sLNvdorsal projection development and suggest that PDF expression in LNvcell bodies and their remaining projections mediate anticipation of the lights-on and lights-off transitions during a light/dark cycle. SIGNIFICANCE STATEMENT: In animals, circadian clocks drive daily rhythms in physiology, metabolism, and behavior via transcriptional feedback loops. Because key circadian transcriptional activators and repressors are regulated by phosphorylation, we screened for phosphatases that alter activity rhythms when their expression was reduced. One such phosphatase, leukocyte-antigen-related (LAR), abolishes activity rhythms, but does not disrupt feedback loop function. Rather,Lardisrupts clock output by eliminating axonal processes from clock neurons that release pigment-dispersing factor (PDF) neuropeptide into the dorsal brain, but PDF expression persists in their cell bodies and remaining projections. In contrast to flies that lack PDF, flies that lackLaranticipate lights-on and lights-off transitions normally, which suggests that the remaining PDF expression mediates activity during light/dark cycles.

Receptor tyrosine phosphatase CLR-1 acts in skin cells to promote sensory dendrite outgrowth.

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.

Chondroitin sulfates and their binding molecules in the central nervous system.

Chondroitin sulfate (CS) is the most abundant glycosaminoglycan (GAG) in the central nervous system (CNS) matrix. Its sulfation and epimerization patterns give rise to different forms of CS, which enables it to interact specifically and with a significant affinity with various signalling molecules in the matrix including growth factors, receptors and guidance molecules. These interactions control numerous biological and pathological processes, during development and in adulthood. In this review, we describe the specific interactions of different families of proteins involved in various physiological and cognitive mechanisms with CSs in CNS matrix. A better understanding of these interactions could promote a development of inhibitors to treat neurodegenerative diseases.

Catalytic and substrate promiscuity: distinct multiple chemistries catalysed by the phosphatase domain of receptor protein tyrosine phosphatase.

The presence of latent activities in enzymes is posited to underlie the natural evolution of new catalytic functions. However, the prevalence and extent of such substrate and catalytic ambiguity in evolved enzymes is difficult to address experimentally given the order-of-magnitude difference in the activities for native and, sometimes, promiscuous substrate/s. Further, such latent functions are of special interest when the activities concerned do not fall into the domain of substrate promiscuity. In the present study, we show a special case of such latent enzyme activity by demonstrating the presence of two mechanistically distinct reactions catalysed by the catalytic domain of receptor protein tyrosine phosphatase isoform δ (PTPRδ). The primary catalytic activity involves the hydrolysis of a phosphomonoester bond (C─O─P) with high catalytic efficiency, whereas the secondary activity is the hydrolysis of a glycosidic bond (C─O─C) with poorer catalytic efficiency. This enzyme also displays substrate promiscuity by hydrolysing diester bonds while being highly discriminative for its monoester substrates. To confirm these activities, we also demonstrated their presence on the catalytic domain of protein tyrosine phosphatase Ω (PTPRΩ), a homologue of PTPRδ. Studies on the rate, metal-ion dependence, pH dependence and inhibition of the respective activities showed that they are markedly different. This is the first study that demonstrates a novel sugar hydrolase and diesterase activity for the phosphatase domain (PD) of PTPRδ and PTPRΩ. This work has significant implications for both understanding the evolution of enzymatic activity and the possible physiological role of this new chemistry. Our findings suggest that the genome might harbour a wealth of such alternative latent enzyme activities in the same protein domain that renders our knowledge of metabolic networks incomplete.

Regulation of receptor-type protein tyrosine phosphatases by their C-terminal tail domains.

Protein tyrosine phosphatases (PTPs) perform specific functions in vivo, despite being vastly outnumbered by their substrates. Because of this and due to the central roles PTPs play in regulating cellular function, PTP activity is regulated by a large variety of molecular mechanisms. We review evidence that indicates that the divergent C-terminal tail sequences (C-terminal domains, CTDs) of receptor-type PTPs (RPTPs) help regulate RPTP function by controlling intermolecular associations in a way that is itself subject to physiological regulation. We propose that the CTD of each RPTP defines an 'interaction code' that helps determine molecules it will interact with under various physiological conditions, thus helping to regulate and diversify PTP function.

Structure-function analyses of tyrosine phosphatase PTP69D in giant fiber synapse formation of Drosophila.

PTP69D is a receptor protein tyrosine phosphatase (RPTP) with two intracellular catalytic domains (Cat1 and Cat2) and has been shown to play a role in axon guidance of embryonic motoneurons as well as targeting of photoreceptor neurons in the visual system of Drosophila melanogaster. Here, we characterized the developmental role of PTP69D in the giant fiber (GF) neurons, two interneurons in the central nervous system (CNS) that control the escape response of the fly. Our studies revealed that PTP69D has a function in synaptic terminal growth in the CNS. We found that missense mutations in the first immunoglobulin (Ig) domain and in the Cat1 domain, present in Ptp69D10 and Ptp69D20 mutants, respectively, did not affect axon guidance or targeting but resulted in stunted terminal growth of the GFs. Cell autonomous rescue experiments demonstrated a function for the Cat1 and the first Ig domain of PTP69D in the GFs but not in its postsynaptic target neurons. In addition, complementation studies and structure-function analyses revealed that for GF terminal growth Cat1 function of PTP69D requires the immunoglobulin and the Cat2 domains, but not the fibronectin III or the membrane proximal region domains. In contrast, the fibronectin III but not the immunoglobulin domains were previously shown to be essential for axon targeting of photoreceptor neurons. Thus, our studies uncover a novel role for PTP69D in synaptic terminal growth in the CNS that is mechanistically distinct from its function in photoreceptor targeting.