The transmembrane protein Off-track 2 is implicated in the guidance of embryonic motor neurons

An essential component of nervous system development entails the projection of axons from neuronal cell bodies to other cells, including other neurons and muscles, resulting in functional circuitry. Drosophila have been indispensible in identifying the proteins underlying axon guidance, which depends on receptors on the surface of growth cones detecting attractive and repulsive ligands. A substantial proportion of this research has utilized embryonic motor neurons, which, in numbering just ~40 per hemisegment, constitute a particularly intelligible system. Bioinformatic analyses of the Drosophila proteome have revealed uncharacterized proteins that might be implicated in axon guidance. These possess the same domains as established molecules and are localized to the cell surface. Proteins meeting these criteria that are also paralogous to established axon guidance molecules and that are expressed in similar spatiotemporal patterns are especially promising candidates. Off-track 2, the focus of the current study, is one such candidate, which has recently been shown to co-precipitate with the putative axon guidance molecule, Off-track. The excision of a P-element located 28 bp upstream of the 5’ UTR of off-track 2 resulted in the appearance of several nucleotides within the promoter region that differ from those of the parent or wild type sequences. Immunohistochemistry revealed that embryos of this line exhibit highly penetrant phenotypes within a number of motor neuron branches; most notably, the ISNb fails to defasciculate from the ISN in ~75% of hemisegments, though several other branches, including the FB and SB, are absent almost to the same extent. The phenotypes of this line mirror those of embryos with a deficiency spanning the gene, suggesting the altered nucleotides resulted in a loss-of-function, presumably by disrupting transcription. Driving Off-track 2 in somatic musculature caused stalling of the ISNb at various choice points, resulting in reduced innervation of the ventral lateral muscles. While these findings suggest Off-track 2 contributes to the pathfinding behavior of embryonic motor neurons, ongoing work is focused on determining the precise role of Off-track 2 in axon guidance

The extracellular Leucine-Rich Repeat superfamily; a comparative survey and analysis of evolutionary relationships and expression patterns

Correction to Dolan J, Walshe K, Alsbury S, Hokamp K, O'Keeffe S, Okafuji T, Miller SF, Tear G, Mitchell KJ: The extracellular leucine-rich repeat superfamily; a comparative survey and analysis of evolutionary relationships and expression patterns. BMC Genomics 2007, 8:320

Neuroprotection and ageing in sensory neurons

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

A bioinformatic and in situ screen for novel axon guidance molecules

Previous screens for axon guidance molecules have identified that many of the molecular cues and their axonal receptors fall into a few major classes that are conserved from invertebrates to vertebrates. While it has been speculated that the majority of axon guidance molecules have already been discovered it seems likely that many such molecules would have been missed due to experimental biases in the genetic or biochemical methods used to identify them. Additionally it seems unlikely that sufficient molecules have been identified to encode the complete wiring of the embryonic nervous system. In order to identify further axon guidance molecules we have taken a systematic bioinformatic approach to identify novel transmembrane proteins that contain any of a number of motifs previously found in known axon guidance molecules. These axon guidance motifs include, for example, immunoglobulin (Ig) motifs, fibronectin-type III (FN3) motifs, leucine-rich repeats (LRR) and epidermal growth factor (EGF) repeats amongst others. This screen has identified 162 genes in Drosophila that fulfil these criteria and their expression patterns were subsequently determined by in situ hybridization. This study yielded 41 candidate axon guidance molecules that show neural expression in the embryo during the period of axon extension. These include 9 genes that appear to have orthologues in vertebrates including the CG32635/Neto and Ten/Odz families. We are now carrying out functional analyses to assess the involvement of these genes in axon guidance in the embryo

Mushroom body defect is required in parallel to Netrin for midline axon guidance in Drosophila

The outgrowth of many neurons within the central nervous system is initially directed towards or away from the cells lying at the midline. Recent genetic evidence suggests that a simple model of differential sensitivity to the conserved Netrin attractants and Slit repellents is insufficient to explain the guidance of all axons at the midline. In the Drosophila embryonic ventral nerve cord, many axons still cross the midline in the absence of the Netrin genes (NetA and NetB) or their receptor frazzled. Here we show that mutation of mushroom body defect (mud) dramatically enhances the phenotype of Netrin or frazzled mutants, resulting in many more axons failing to cross the midline, although mutations in mud alone have little effect. This suggests that mud, which encodes a microtubule-binding coiled-coil protein homologous to NuMA and LIN-5, is an essential component of a Netrin-independent pathway that acts in parallel to promote midline crossing. We demonstrate that this novel role of Mud in axon guidance is independent of its previously described role in neural precursor development. These studies identify a parallel pathway controlling midline guidance in Drosophila and highlight a novel role for Mud potentially acting downstream of Frizzled to aid axon guidance

The extracellular Leucine-Rich Repeat superfamily; a comparative survey and analysis of evolutionary relationships and expression patterns

BACKGROUND: Leucine-rich repeats (LRRs) are highly versatile and evolvable protein-ligand interaction motifs found in a large number of proteins with diverse functions, including innate immunity and nervous system development. Here we catalogue all of the extracellular LRR (eLRR) proteins in worms, flies, mice and humans. We use convergent evidence from several transmembrane-prediction and motif-detection programs, including a customised algorithm, LRRscan, to identify eLRR proteins, and a hierarchical clustering method based on TribeMCL to establish their evolutionary relationships. RESULTS: This yields a total of 369 proteins (29 in worm, 66 in fly, 135 in mouse and 139 in human), many of them of unknown function. We group eLRR proteins into several classes: those with only LRRs, those that cluster with Toll-like receptors (Tlrs), those with immunoglobulin or fibronectin-type 3 (FN3) domains and those with some other domain. These groups show differential patterns of expansion and diversification across species. Our analyses reveal several clusters of novel genes, including two Elfn genes, encoding transmembrane proteins with eLRRs and an FN3 domain, and six genes encoding transmembrane proteins with eLRRs only (the Elron cluster). Many of these are expressed in discrete patterns in the developing mouse brain, notably in the thalamus and cortex. We have also identified a number of novel fly eLRR proteins with discrete expression in the embryonic nervous system. CONCLUSION: This study provides the necessary foundation for a systematic analysis of the functions of this class of genes, which are likely to include prominently innate immunity, inflammation and neural development, especially the specification of neuronal connectivity