S. Lawrence Zipursky

Drosophila Dscam Is an Axon Guidance Receptor Exhibiting Extraordinary Molecular Diversity

A Drosophila homolog of human Down syndrome cell adhesion molecule (DSCAM), an immunoglobulin superfamily member, was isolated by its affinity to Dock, an SH3/SH2 adaptor protein required for axon guidance. Dscam binds directly to both Docks SH2 and SH3 domains. Genetic studies revealed that Dscam, Dock and Pak, a serine/threonine kinase, act together to direct pathfinding of Bolwigs nerve, containing a subclass of sensory axons, to an intermediate target in the embryo. Dscam also is required for the formation of axon pathways in the embryonic central nervous system. cDNA and genomic analyses reveal the existence of multiple forms of Dscam with a conserved architecture containing variable Ig and transmembrane domains. Alternative splicing can potentially generate more than 38,000 Dscam isoforms. This molecular diversity may contribute to the specificity of neuronal connectivity.

The Eye-Specification Proteins So and Eya Form a Complex and Regulate Multiple Steps in Drosophila Eye Development

Sine oculis (so) and eyes absent (eya) are required for Drosophila eye development and are founding members of the mammalian Six and Eya gene families. These genes have been proposed to act with eyeless (Pax6) to regulate eye development in vertebrates and invertebrates. so encodes a highly diverged homeobox transcription factor and eya encodes a novel nuclear protein. We demonstrate that So and Eya (1) regulate common steps in eye development including cell proliferation, patterning, and neuronal development; (2) synergize in inducing ectopic eyes; and (3) interact in yeast and in vitro through evolutionarily conserved domains. We propose that an So/Eya complex regulates multiple steps in eye development and functions within the context of a network of genes to specify eye tissue identity.

Polyglutamine-expanded human huntingtin transgenes induce degeneration of Drosophila photoreceptor neurons

Huntingtons disease (HD) is an autosomal dominant neurodegenerative disorder. Disease alleles contain a trinucleotide repeat expansion of variable length, which encodes polyglutamine tracts near the amino terminus of the HD protein, huntingtin. Polyglutamine-expanded huntingtin, but not normal huntingtin, forms nuclear inclusions. We describe a Drosophila model for HD. Amino-terminal fragments of human huntingtin containing tracts of 2, 75, and 120 glutamine residues were expressed in photoreceptor neurons in the compound eye. As in human neurons, polyglutamine-expanded huntingtin induced neuronal degeneration. The age of onset and severity of neuronal degeneration correlated with repeat length, and nuclear localization of huntingtin presaged neuronal degeneration. In contrast to other cell death paradigms in Drosophila, coexpression of the viral antiapoptotic protein, P35, did not rescue the cell death phenotype induced by polyglutamine-expanded huntingtin.

The drosophila sine oculis locus encodes a homeodomain-containing protein required for the development of the entire visual system

The transformation of an unpatterned epithelium into a patterned one is a fundamental issue in morphogenesis. This transformation occurs in a dramatic fashion in the developing eye imaginal disc, the primordium of the Drosophila compound eye. Molecular and developmental analyses reveals that the sine oculis (so) locus encodes a homeodomain-containing protein that is expressed and required in the unpatterned epithelium prior to morphogenesis. In mutants, cells undergo apoptosis. These findings argue that so plays an essential role in controlling the initial events of pattern formation in the eye disc. So is also expressed and required for the development of the rest of the fly visual system, including the optic lobes (i.e., those regions of the brain that process visual information). So is expressed in the optic lobe primordium prior to its invagination from the embryonic ectoderm; in so mutants, the optic lobe primordium fails to invaginate.

A gain-of-function mutation in Drosophila MAP kinase activates multiple receptor tyrosine kinase signaling pathways

In the Drosophila eye, activation of the sevenless (sev) receptor tyrosine kinase is required for the specification of the R7 photoreceptor cell fate. In a genetic screen for mutations that result in the activation of the sev signaling pathway in the absence of the inducing signal, we identified a gain-of-function mutation in rolled (rlSevenmaker [rlSem]), which encodes a homolog of mitogen-activated protein (MAP) kinase. In addition to the sev pathway, this mutation activates the pathways controlled by torso and the epidermal growth factor receptor homology. The rlSem mutation results in the substitution of a single conserved amino acid in the kinase domain. Activation of MAP kinase by the rlSem mutation is both necessary and sufficient to activate multiple signaling pathways controlled by receptor tyrosine kinases.

DESIGN PRINCIPLES OF INSECT AND VERTEBRATE VISUAL SYSTEMS

A century ago, Cajal noted striking similarities between the neural circuits that underlie vision in vertebrates and flies. Over the past few decades, structural and functional studies have provided strong support for Cajals view. In parallel, genetic studies have revealed some common molecular mechanisms controlling development of vertebrate and fly visual systems and suggested that they share a common evolutionary origin. Here, we review these shared features, focusing on the first several layers-retina, optic tectum (superior colliculus), and lateral geniculate nucleus in vertebrates; and retina, lamina, and medulla in fly. We argue that vertebrate and fly visual circuits utilize common design principles and that taking advantage of this phylogenetic conservation will speed progress in elucidating both functional strategies and developmental mechanisms, as has already occurred in other areas of neurobiology ranging from electrical signaling and synaptic plasticity to neurogenesis and axon guidance.

Pak functions downstream of Dock to regulate photoreceptor axon guidance in Drosophila.

The SH2/SH3 adaptor protein Dock has been proposed to transduce signals from guidance receptors to the actin cytoskeleton in Drosophila photoreceptor (R cell) growth cones. Here, we demonstrate that Drosophila p21-activated kinase (Pak) is required in a Dock pathway regulating R cell axon guidance and targeting. Dock and Pak colocalize to R cell axons and growth cones, physically interact, and their loss-of-function phenotypes are indistinguishable. Normal patterns of R cell connectivity require Paks kinase activity and binding sites for both Dock and Cdc42/Rac. A membrane-tethered form of Pak (Pak(myr) acts as a dominant gain-of-function protein. Retinal expression of Pak(myr) rescues the R cell connectivity phenotype in dock mutants. These data establish Pak as a critical regulator of axon guidance and a downstream effector of Dock in vivo.

Drosophila Photoreceptor Axon Guidance and Targeting Requires the Dreadlocks SH2/SH3 Adapter Protein

SUMMARY Mutations in the Drosophila gene dreadlocks (dock) disrupt photoreceptor cell (R cell) axon guidance and targeting. Genetic mosaic analysis and cell-type-specific expression of dock transgenes demonstrate dock is required in R cells for proper innervation. Dock protein contains one SH2 and three SH3 domains, implicating it in tyrosine kinase signaling, and is highly related to the human proto-oncogene Nck. Dock expression is detected in R cell growth cones in the target region. We propose Dock transmits signals in the growth cone in response to guidance and targeting cues. These findings provide an important step for dissection of signaling pathways regulating growth cone motility.

Alternative Splicing of Drosophila Dscam Generates Axon Guidance Receptors that Exhibit Isoform-Specific Homophilic Binding

Dscam is an immunoglobulin (Ig) superfamily protein required for the formation of neuronal connections in Drosophila. Through alternative splicing, Dscam potentially gives rise to 19,008 different extracellular domains linked to one of two alternative transmembrane segments, resulting in 38,016 isoforms. All isoforms share the same domain structure but contain variable amino acid sequences within three Ig domains in the extracellular region. We demonstrate that different isoforms exhibit different binding specificity. Each isoform binds to itself but does not bind or binds poorly to other isoforms. The amino acid sequences of all three variable Ig domains determine binding specificity. Even closely related isoforms sharing nearly identical amino acid sequences exhibit isoform-specific binding. We propose that this preferential homophilic binding specificity regulates interactions between cells and contributes to the formation of complex patterns of neuronal connections.

N-Cadherin Regulates Target Specificity in the Drosophila Visual System

Using visual behavioral screens in Drosophila, we identified multiple alleles of N-cadherin. Removal of N-cadherin selectively from photoreceptor neurons (R cells) causes deficits in specific visual behaviors that correlate with disruptions in R cell connectivity. These defects include disruptions in the pattern of neuronal connections made by all three classes of R cells (R1-R6, R7, and R8). N-cadherin is expressed in both R cell axons and their targets. By inducing mitotic recombination in a subclass of eye progenitors, we generated mutant R7 axons surrounded by largely wild-type R cell axons and a wild-type target. R7 axons lacking N-cadherin mistarget to the R8 recipient layer. We consider the implications of these findings in the context of the proposed role for cadherins in target specificity.