Activity-Independent Prespecification of Synaptic Partners in the Visual Map of Drosophila

SummarySpecifying synaptic partners and regulating synaptic numbers are at least partly activity-dependent processes during visual map formation in all systems investigated to date [1–5]. In Drosophila, six photoreceptors that view the same point in visual space have to be sorted into synaptic modules called cartridges in order to form a visuotopically correct map [6, 7]. Synapse numbers per photoreceptor terminal and cartridge are both precisely regulated [8–10]. However, it is unknown whether an activity-dependent mechanism or a genetically encoded developmental program regulates synapse numbers. We performed a large-scale quantitative ultrastructural analysis of photoreceptor synapses in mutants affecting the generation of electrical potentials (norpA, trp;trpl), neurotransmitter release (hdc, syt), vesicle endocytosis (synj), the trafficking of specific guidance molecules during photoreceptor targeting (sec15), a specific guidance receptor required for visual map formation (Dlar), and 57 other novel synaptic mutants affecting 43 genes. Remarkably, in all these mutants, individual photoreceptors form the correct number of synapses per presynaptic terminal independently of cartridge composition. Hence, our data show that each photoreceptor forms a precise and constant number of afferent synapses independently of neuronal activity and partner accuracy. Our data suggest cell-autonomous control of synapse numbers as part of a developmental program of activity-independent steps that lead to a “hard-wired” visual map in the fly brain

Weg- und Zielfindung wachsender Nervenfasern im Sehsystem von Fliegen und Fischen

Der Vergleich der Weg- und Zielfindung von Sehfasern bei Fliegen und Fischen ist attraktiv, da die wachsenden Nervenfasern trotz der verschiedenen Augenformen möglicherweise vergleichbare Strategien verwenden. Es ist sogar möglich, daß die grundlegenden Mechanismen der Zellerkennung bereits so früh in der Evolution erworben wurden, daß homologe molekulare Komponenten bei Vertebraten und Invertebraten hieran beteiligt sind

The Cell Adhesion Molecules Roughest, Hibris, Kin of Irre and Sticks and Stones Are Required for Long Range Spacing of the Drosophila Wing Disc Sensory Sensilla

Most animal tissues and organ systems are comprised of highly ordered arrays of varying cell types. The development of external sensory organs requires complex cell-cell communication in order to give each cell a specific identity and to ensure a regular distributed pattern of the sensory bristles. This involves both long and short range signaling mediated by either diffusible or cell anchored factors. In a variety of processes the heterophilic Irre Cell Recognition Module, consisting of the Neph-like proteins: Roughest, Kin of irre and of the Nephrin-like proteins: Sticks and Stones, Hibris, plays key roles in the recognition events of different cell types throughout development. In the present study these proteins are apically expressed in the adhesive belt of epithelial cells participating in sense organ development in a partially exclusive and asymmetric manner. Using mutant analysis the GAL4/UAS system, RNAi and gain of function we found an involvement of all four Irre Cell Recognition Module-proteins in the development of a highly structured array of sensory organs in the wing disc. The proteins secure the regular spacing of sensory organs showing partial redundancy and may function in early lateral inhibition events as well as in cell sorting processes. Comparisons with other systems suggest that the Irre Cell Recognition module is a key organizer of highly repetitive structures.status: publishe

Loss- and gain-of-function analysis of the lipid raft proteins Reggie/Flotillin in Drosophila : they are posttranslationally regulated, and misexpression interferes with wing and eye development

Reggie/Flotillin proteins are upregulated after optic nerve dissection and evolutionary highly conserved components of lipid rafts. Whereas many biochemical and cell culture studies suggest an involvement in the assembly of multiprotein complexes at cell contact sites, not much is known about their biological in vivo functions. We therefore set out to study the expression pattern and the effects of loss- and gain-of-function in the Drosophila melanogaster model system. We found that in flies these proteins are mainly expressed in axons at the root of fiber tracts, in places where strong fasciculation is required, e.g. at the neck of the peduncle of the mushroom bodies and in the optic chiasms. Despite their evolutionary conservation which implies fundamental and important functions, a P-element-induced null mutant (KG00210) of reggie1/flotillin2 (reggie1/flo2) in D. melanogaster shows no apparent phenotypic defects. This was even more surprising as we show that in this reggie1/flo2 null mutant the paralogous Reggie2/Flo1 protein is unstable and degraded, while the transcript is still present. The requirement of Reggie1/Flo2 for Reggie2/Flo1 stabilization is confirmed by misexpression experiments. Reggie2/Flo1 can only be misexpressed when Reggie1/Flo2 is provided as well. Conversely, Reggie1/Flo2 immunoreactivity can be detected, when its transgene is misexpressed alone. Using appropriate Gal4 driver lines, misexpression of Reggie1/Flo2 alone or together with Reggie2/Flo1 in the eye imaginal disc results in a specific and severe mislocalization of cell adhesion molecules of the immunoglobulin superfamily (IgCAMs) (while DE-Cadherin is unaffected) and in differentiation defects pointing to impaired signaling. In the wing imaginal disc, global overexpression of Reggie/Flotillin proteins leads to a significant extension of the Wingless signal and severely disrupts normal wing development. Our data support the notion that Reggie/Flotillin proteins are implicated in signaling processes at cellular contact sites

Irregular chiasm-C-roughest, a member of the immunoglobulin superfamily, affects sense organ spacing on the Drosophila antenna by influencing the positioning of founder cells on the disc ectoderm

We describe a role for Irregular chiasmC-roughest (IrreC-rst), an immunoglobulin (Ig) superfamily member, in patterning sense organs on the Drosophila antenna. IrreC-rst protein is initially expressed homogeneously on apical profiles of ectodermal cells in regions of the antennal disc. During specification of founder cells (FCs), the intracellular protein distribution changes and becomes concentrated in regions where specific intercellular contacts presumably occur. Loss of function mutations as well as misexpression of irreC-rst results in an altered arrangement of FCs within the disc compared to wildtype. Sense organ development occurs normally, although spacing is affected. Unlike its role in interommatidial spacing, irreC-rst does not affect apoptosis during antennal development. We propose that IrreC-rst affects the spatial relationship between sensory and ectodermal cells during FC delamination

Model of IRM-protein interactions in the <i>Drosophila</i> anterior wing margin.

<p>(A-E) Illustration of IRM-protein functions in the anterior wing margin. In black are epithelial cells shown, while SOPs are shown in orange. Protein interactions are shown in different sizes according to the strength of the interaction. Red represents Rst, green Hbs, blue Kirre and SNS is shown in yellow. In the wild type (A) preferred adhesion was observed between the SOPs and the surrounding epithelial cells. Epithelial cells are additionally stable connected through the Hbs, Rst and Hbs, Kirre interaction. In <i>rst</i>^<i>1R34</i> (B), as an example for Neph-like loss of function, preferential adhesion can still be observed between the SOPs and epithelial cells through the SNS, Kirre and Hbs Kirre interaction. Only in the case of Rst and Kirre loss, the adhesive properties of the wing margin is changed leading to bristle clusters (C). Loss of Hbs prevents heterophilic interaction between the non-SOP cells resulting in mild disturbances of the SOP pattern (D). Loss of Hbs and SNS results in total loss of heterophilic interaction between all cell types in the presumptive anterior wing margin (E). This results in strong disturbances of the SOP and later the bristle pattern (F) Summary of the inductive and competitive interactions between the IRM-proteins <i>in trans</i> and <i>in cis</i>. In the interaction between two cells <i>in trans</i> several inductive events were observed, if these events represent inductions of gene expression or stabilization of proteins in the adhesive belt by heterophilic interactions is currently unknown. Inside cells <i>in cis</i> several competitive interactions were observed, resulting in degradation of proteins in vesicles. Altogether, these interactions allow a precise regulation of IRM-protein abundance and function. (G) Chain model of preferential adhesion of IRM-proteins in the wing disc. The IRM-proteins in the wing disc secure a strong adhesive chain in the distal growing wing. Preferential adhesion around the SOPs secures a constant high number of cells between the SOPs. Growth in distal directions explains the lower number of cells between precursors compared to the adult sensory organs.</p

Hbs acts cooperatively with SNS to secure the bristle pattern in the anterior wing margin.

<p>(A-L) Projection views of IRM-protein immunoreactivity in late third instar larvae. Rst is shown in red (A, E and I), Hbs in green (B, F and N), Kirre in blue (C, G and K) and SNS in yellow (D, H and L). (A-D) Global <i>hbs-RNAi</i> using <i>MZ1369-GAL4</i> reduces the staining for Rst (A) and Kirre (C) in all membranes that are not in contact to the SOPs (B) Hbs immunoreactivity is reduced and no clear membrane localization is detectable. (D) SNS immunoreactivity is only mildly affected, but SOPs stand significantly nearer to each other. (E-H) <i>MZ1369-GAL4</i>><i>UAS-sns-RNAi</i> shows mildly reduced Rst staining (E). Hbs (F) and Kirre (K) immunoreactivity is unchanged. SNS (H) is not detectable. (I-L) In the double RNAi <i>MZ1369>hbs-RNAi</i>, <i>SNS-RNAi</i> only the two adhesive belts with Rst (I) and Kirre (K) are visible, but no obvious SOPs are marked. Hbs (J) and SNS (L) are not detectable. (M) In the adult <i>MZ1369-GAL4</i> driven <i>hbs-RNAi</i> shows only a mild spacing phenotype with 0 to 7 intervening cells. (N) The global <i>sns-RNAi</i> in the entire wing disc shows a very mild spacing phenotype with spacing ranging from 1 to 5. (O) <i>MZ1369>hbs-RNAi</i>, <i>sns-RNAi</i> shows a significant disturbance of the spacing of recurved bristles with spacing ranging from 0 to 8. (P) <i>MZ1369-GAL4</i> driven misexpression of <i>hbs</i> has a strong impact on the spacing of recurved bristles with spacing ranging from 0 to 12. Additionally, clustered recurved bristles are frequently observed. (Q) Quantitative analysis of the recurved bristle spacing, as measured by the number of slender bristles between the recurved bristles. The distribution of <i>MZ1369>GFP</i> differs significantly from <i>MZ1369>hbs-RNAi</i> in the following spacing value: 3: p-value = 0.009. <i>MZ1369>SNS-RNAi</i> differs significantly in the following spacing value: < = 1: p-value = 0.035. The double RNAi for <i>hbs</i> and <i>SNS</i> is significantly different for: < = 1: p-value = 0.0002, 3: p-value = 0.014. <i>hbs</i> overexpression differs for: < = 1: p-value = 0.0011, 3: p-value = 0.0013, > = 6: p-value = <0.001. Scale bars correspond to 10μm in all images.</p