Research Article

Branch-restricted localization of phosphatase Prl-1 specifies axonal synaptogenesis domains

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Science  03 May 2019:
Vol. 364, Issue 6439, eaau9952
DOI: 10.1126/science.aau9952

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Compartmentalization of axons

Neurons can spread branches far and wide across the central nervous system. One neuron can simultaneously contact several other neurons, amplifying network complexity. Studying the Drosophila brain, Urwyler et al. found that not all axon branches are the same, even within one neuron (see the Perspective by Falkner and Scheiffele). A membrane-anchored phosphatase functions within a subset of axon branches to regulate synapse formation. Thus, the pattern of synapses on one neuron is not random but rather is spatially restricted according to subcompartments in the axon.

Science, this issue p. eaau9952; see also p. 437

Structured Abstract

INTRODUCTION

Axons grow collateral branches to project to distinct target areas in the central nervous system (CNS) and to form synapses with various postsynaptic partner cells. Synapse numbers, locations, and types can differ in different axon collaterals of the same neuron and need to be specified independently in each collateral. Which mechanisms control synapse formation with subcellular and spatial specificity in the CNS? Signaling dependent on cell surface receptors is essential in the processes of axon guidance, branching, and synaptogenesis. However, how intracellular axon-intrinsic factors control compartment-specific synaptogenesis in the CNS remains unclear.

RATIONALE

Using a genetic single-cell approach that allows the labeling and manipulation of single, specific mechanosensory axons in the Drosophila CNS, we searched for molecular players controlling synapse formation specifically in one subcellular compartment and target area. Protein kinases and phosphatases control reversible phosphorylation cascades that are central to most intracellular signaling pathways. We reasoned that these kinases and phosphatases are likely also key to controlling the spatial specificity of synapse formation and screened the fly kinome and phosphatome by an in vivo cell-autonomous knockdown approach.

RESULTS

We found that the loss of phosphatase of regenerating liver (Prl-1) specifically reduces synapses organized in a terminal arbor in one target area of mechanosensory neurons. Collaterals that target other CNS regions do not display axonal or synaptic defects. Prl family members are small, membrane-localized phosphatases conserved from invertebrates to mammals. They are associated with metastatic progression of tumors. Prl genes are also expressed in the CNSs of flies and mice; however, their neuronal functions are not known. In this study, we show that the loss of Drosophila Prl-1 leads to defects in axonal target areas in several CNS circuits. The CNS of prl-1 null mutant flies is reduced in size, and the animals have locomotor defects. A developmental analysis in mechanosensory neurons revealed that Prl-1 is required for the stabilization of nascent axonal arbors and synaptic structures. Prl-1 overexpression induces ectopic axonal protrusions and synapses. Prl proteins are dual-specificity phosphatases, and human Prl-3 was suggested to dephosphorylate not only protein substrates but also the phosphoinositide phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. We provide genetic evidence consistent with the model that Prl-1 leads to the reduction of PI(4,5)P2 in the process of local synaptogenesis.

Moreover, our data show that the synaptogenic function of Prl-1 involves axon branch–specific modulation of the insulin receptor (InR) signaling pathway. Reduction of InR, phosphatidylinositol 3-kinase, Akt, or Raptor, presynaptically, phenocopies the branch-specific loss of synaptic arborizations. Conversely, the knockdown of PTEN or the expression of constitutively active Akt substantially increases the number of presynapses, an effect that can be suppressed by the loss of Prl-1. Therefore, inactivation of this signaling cascade produces the same spatially restricted synaptic defects as the loss of Prl-1. Lastly, we show that Prl-1 protein gets selectively localized to the axon compartment in which its function is required. Compartment-specific localization and function of Prl-1 depend on long untranslated sequences in the prl-1 mRNA.

CONCLUSION

Prl phosphatase regulates CNS circuit formation. Prl-1 modulates InR-Akt signaling, likely by targeting membrane phosphoinositides, to control synapse formation specifically in one axon collateral of mechanosensory neurons. We suggest that untranslated prl-1 mRNA elements could mediate the local translation of Prl-1 in axonal subcompartments. Thus, Prl-1 could provide a specificity factor to restrict Akt signaling and synapse formation in a subcellular compartment of neurons.

Axon branch–specific localization of Prl-1 directs CNS synaptogenesis.

Mechanosensory axons innervate distinct CNS target areas by forming three primary collaterals (1 to 3). Each collateral assembles specific types and numbers of synapses. Prl-1 phosphatase and the InR-Akt signaling pathway are specifically required for the formation of terminal arbor synapses in the contralateral collateral (2), and exuberant synaptogenesis is induced by activated Akt in this collateral. The loss of Prl-1 can be rescued by active Akt. Prl-1 protein is enriched in axon collateral 2, thereby providing spatial specificity. PI3K, phosphatidylinositol 3-kinase.

Abstract

Central nervous system (CNS) circuit development requires subcellular control of synapse formation and patterning of synapse abundance. We identified the Drosophila membrane-anchored phosphatase of regenerating liver (Prl-1) as an axon-intrinsic factor that promotes synapse formation in a spatially restricted fashion. The loss of Prl-1 in mechanosensory neurons reduced the number of CNS presynapses localized on a single axon collateral and organized as a terminal arbor. Flies lacking all Prl-1 protein had locomotor defects. The overexpression of Prl-1 induced ectopic synapses. In mechanosensory neurons, Prl-1 modulates the insulin receptor (InR) signaling pathway within a single contralateral axon compartment, thereby affecting the number of synapses. The axon branch–specific localization and function of Prl-1 depend on untranslated regions of the prl-1 messenger RNA (mRNA). Therefore, compartmentalized restriction of Prl-1 serves as a specificity factor for the subcellular control of axonal synaptogenesis.

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