Liprin-␣ proteins are adaptors that interact with the receptor protein tyrosine phosphatase leukocyte common antigen-related (LAR) and other synaptic proteins to promote synaptic partner selection and active zone assembly. Liprin-␤ proteins bind to and share homology with Liprin-␣ proteins, but their functions at the synapse are unknown. The Drosophila genome encodes single Liprin-␣ and Liprin-␤ homologs, as well as a third related protein that we named Liprin-␥. We show that both Liprin-␤ and Liprin-␥ physically interact with Liprin-␣ and that Liprin-␥ also binds to LAR. Liprin-␣ mutations have been shown to disrupt synaptic target layer selection by R7 photoreceptors and to reduce the size of larval neuromuscular synapses. We have generated null mutations in Liprin-␤ and Liprin-␥ to investigate their role in these processes. We find that, although Liprin-␣ mutant R7 axons terminate before reaching the correct target layer, Liprin-␤ mutant R7 axons grow beyond their target layer. Larval neuromuscular junction size is reduced in both Liprin-␣ and Liprin-␤ mutants, and further reduced in double mutants, suggesting independent functions for these Liprins. Genetic interactions demonstrate that both Liprin proteins act through the exchange factor Trio to promote stable target selection by R7 photoreceptor axons and growth of neuromuscular synapses. Photoreceptor and neuromuscular synapses develop normally in Liprin-␥ mutants; however, removing Liprin-␥ improves R7 targeting in Liprin-␣ mutants, and restores normal neuromuscular junction size to Liprin-␤ mutants, suggesting that Liprin-␥ counteracts the functions of the other two Liprins. We propose that context-dependent interactions between the three Liprins modulate their functions in synapse formation.

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-␣ gene based on a similar R7 projection defect. Liprin-␣ 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-␣ nor the LAR D2 domain. Although human and Caenorhabditis elegans Liprin-␣ 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-␣. In addition, Liprin-␣ overexpression or loss of function can affect R7 targeting in the complete absence of LAR. We conclude that Liprin-␣ does not simply act by regulating LAR localization but also has LAR-independent functions. receptor tyrosine phosphatase ͉ visual system ͉ Drosophila T he color-sensitive photoreceptors of the Drosophila visual system, R7 and R8, provide a simple system in which to study layer-specific axon targeting. Whereas the outer photoreceptors R1-R6 project their axons to the lamina, R7 and R8 project to the medulla, where R8 terminates in the more superficial M3 layer and R7 in the deeper M6 layer. This targeting occurs in two stages, with both R7 and R8 growth cones pausing in separate temporary layers before proceeding to their final positions (1). Several genes are known to contribute to the establishment of the R7 and R8 projection pattern. The transcription factor Runt is expressed in R7 and R8, and its misexpression is sufficient to target R2 and R5 to the medulla, suggesting that it controls the choice of optic neuropil (2). Endogenous expression of the homophilic cell adhesion molecule Capricious (Caps) in R8 or its ectopic expression in R7 directs these photoreceptors to terminate in the Caps-positive M3 layer (3). The transmembrane cadherin Flamingo (Fmi) is required for R8 targeting (4, 5), whereas loss of either N-cadherin (Ncad) (6) or one of the receptor protein tyrosine phosphatases (RPTPs), PTP69D (7) or LAR (8, 9), causes R7 to terminate inappropriately in the R8 layer.Other functions of LAR include axonal patterning of photoreceptors R1-R6 (9) and embryonic motor neurons (10, 11), synapse morphogenesis at the larval neuromuscular junction (NMJ) (12), and polarization of actin filaments in the follicle cells surrounding the oocyte, which promotes egg elongation along the anterior-posterior axis (13,14). It is unclear how LAR and other RPTPs signal within the cell to induce the cytoskeletal rearrangements that mediate these functions. Trio, a guanine nucleotide exchange factor for Rac (15), and Enabled (Ena), which regulates actin polymerization (16), show genetic interactions with LAR in both R7 targeting (8) and motor axon guidance ...

Receptor protein tyrosine phosphatases (RPTPs) control many aspects of nervous system development. At the Drosophila neuromuscular junction (NMJ), regulation of synapse growth and maturation by the RPTP LAR depends on catalytic phosphatase activity and on the extracellular ligands Syndecan and Dally-like. We show here that the function of LAR in controlling R7 photoreceptor axon targeting in the visual system differs in several respects. The extracellular domain of LAR important for this process is distinct from the domains known to bind Syndecan and Dally-like, suggesting the involvement of a different ligand. R7 targeting does not require LAR phosphatase activity, but instead depends on the phosphatase activity of another RPTP, PTP69D. In addition, a mutation that prevents dimerization of the intracellular domain of LAR interferes with its ability to promote R7 targeting, although it does not disrupt phosphatase activity or neuromuscular synapse growth. We propose that LAR function in R7 is independent of its phosphatase activity, but requires structural features that allow dimerization and may promote the assembly of downstream effectors.synapse ͉ neuromuscular junctions ͉ dimerization ͉ wedge R eceptor protein tyrosine phosphatases (RPTPs) are required for nervous system development in both vertebrates and invertebrates (1). Of the six Drosophila RPTPs, Leukocyte antigen-related (LAR) has been studied in most detail due to its non-redundant role in several developmental processes. In Lar mutant embryos, motor neurons in the intersegmental nerve b (ISNb) fail to innervate the appropriate muscles and aberrantly track along the ISN (2). LAR has two distinct functions at the synapses formed by larval motor neurons on their target muscles. Synapse size as defined by the number of synaptic boutons present at these larval neuromuscular junctions (NMJs) is proportional to Lar dosage; and LAR controls active zone morphogenesis and thus synaptic strength (3). In the visual system, LAR enables photoreceptor axons to establish connections to the correct synaptic partners. Photoreceptors R1-R6 project into the lamina, where the axons from a single ommatidium defasciculate and connect to six different laminar cartridges; this defasciculation requires Lar (4). Photoreceptors R7 and R8, which mediate color vision, project beyond the lamina to terminate in two distinct layers of the medulla, R8 in M3 and R7 in the deeper M6 layer (5). In Lar mutants, most R7 axons terminate inappropriately in M3, the same layer as R8 (4, 6).LAR and its vertebrate homologues PTP and PTP␦ are type IIa RPTPs, which have two intracellular phosphatase domains (D1 and D2) and extracellular Ig (Ig) and fibronectin type III (FNIII) domains. The membrane-distal D2 domains of such RPTPs show no phosphatase activity on artificial substrates in vitro (7-9). Nevertheless, the LAR D2 domain is essential for R7 targeting, where it may act by recruiting the scaffolding protein Liprin-␣ (10) or regulating the activity of the D1 domain (8). An important class...

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In the Drosophila adult visual system, photoreceptor axons and their connecting interneurons are tied into a retinotopic pattern throughout the consecutive neuropil regions: lamina, medulla and lobula complex. Lamina and medulla are joined by the first or outer optic chiasm (OOC). Medulla, lobula and lobula plate are connected by the second or inner optic chiasm (IOC). In the regulatory mutant In(1)omb(H31) of the T-box gene optomotor-blind (omb), fibers were found to cross aberrantly through the IOC into the neuropil of the lobula complex. Here, we show that In(1)omb(H31) causes selective loss of OMB expression from glial cells within the IOC previously identified as IOC giant glia (ICg-glia). In the absence of OMB, ICg-glia retain their glial cell identity and survive until the adult stage but tend to be displaced into the lobula complex neuropil leading to a misprojection of axons through the IOC. In addition, adult mutant glia show an aberrant increase in length and frequency of glial cell processes. We narrowed down the onset of the IOC defect to the interval between 48 h and 72 h of pupal development. Within the 40 kb of regulatory DNA lacking in In(1)omb(H31), we identified an enhancer element (ombC) with activity in the ICg-glia. ombC-driven expression of omb in ICg-glia restored proper axonal projection through the IOC in In(1)omb(H31) mutant flies, as well as proper glial cell positioning and morphology. These results indicate that expression of the transcription factor OMB in ICg-glial cells is autonomously required for glial cell migration and morphology and non-autonomously influences axonal pathfinding.