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Contractile actomyosin and protrusive branched F-actin networks interact in a dynamic balance, repeatedly contracting and expanding apical cell contacts to organize the epithelium of the developing fly retina. Previously we showed that the immunoglobulin superfamily protein Sidekick (Sdk) contributes to contraction by recruiting the actin binding protein Polychaetoid (Pyd) to vertices. Here we show that as tension increases during contraction, Sdk progressively accumulates at vertices, where it toggles to recruit the WAVE regulatory complex (WRC) to promote actin branching and protrusion. Sdk alternately interacts with the WRC and Pyd using the same C-terminal motif. With increasing protrusion, levels of Sdk and the WRC decrease at vertices while levels of Pyd increase paving the way for another round of contraction. Thus, by virtue of dynamic association with vertices and interchangeable associations with contractile and protrusive effectors, Sdk is central to controlling the balance between contraction and expansion that shapes this epithelium., (© 2022 Malin et al.)

Tricellular adherens junctions are points of high tension that are central to the rearrangement of epithelial cells. However, the molecular composition of these junctions is unknown, making it difficult to assess their role in morphogenesis. Here, we show that Sidekick, an immunoglobulin family cell adhesion protein, is highly enriched at tricellular adherens junctions in Drosophila. This localization is modulated by tension, and Sidekick is itself necessary to maintain normal levels of cell bond tension. Loss of Sidekick causes defects in cell and junctional rearrangements in actively remodeling epithelial tissues like the retina and tracheal system. The adaptor proteins Polychaetoid and Canoe are enriched at tricellular adherens junctions in a Sidekick-dependent manner; Sidekick functionally interacts with both proteins and directly binds to Polychaetoid. We suggest that Polychaetoid and Canoe link Sidekick to the actin cytoskeleton to enable tricellular adherens junctions to maintain or transmit cell bond tension during epithelial cell rearrangements., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The assembly of functional neuronal circuits requires growth cones to extend in defined directions and recognize the correct synaptic partners. Homophilic adhesion between vertebrate Sidekick proteins promotes synapse formation between retinal neurons involved in visual motion detection. We show here that Drosophila Sidekick accumulates in specific synaptic layers of the developing motion detection circuit and is necessary for normal optomotor behavior. Sidekick is required in photoreceptors, but not in their target lamina neurons, to promote the alignment of lamina neurons into columns and subsequent sorting of photoreceptor axons into synaptic modules based on their precise spatial orientation. Sidekick is also localized to the dendrites of the direction-selective T4 and T5 cells, and is expressed in some of their presynaptic partners. In contrast to its vertebrate homologs, Sidekick is not essential for T4 and T5 to direct their dendrites to the appropriate layers or to receive synaptic contacts. These results illustrate a conserved requirement for Sidekick proteins in establishing visual motion detection circuits that is achieved through distinct cellular mechanisms in Drosophila and vertebrates., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Wnt target gene transcription is mediated by nuclear translocation of stabilized beta-catenin, which binds to TCF and recruits Pygopus, a cofactor with an unknown mechanism of action. The mediator complex is essential for the transcription of RNA polymerase II-dependent genes; it associates with an accessory subcomplex consisting of the Med12, Med13, Cdk8, and Cyclin C subunits. We show here that the Med12 and Med13 subunits of the Drosophila mediator complex, encoded by kohtalo and skuld, are essential for the transcription of Wingless target genes. kohtalo and skuld act downstream of beta-catenin stabilization both in vivo and in cell culture. They are required for transcriptional activation by the N-terminal domain of Pygopus, and their physical interaction with Pygopus depends on this domain. We propose that Pygopus promotes Wnt target gene transcription by recruiting the mediator complex through interactions with Med12 and Med13.

The organizing centers for Drosophila imaginal disc development are created at straight boundaries between compartments; these are maintained by differences in cell affinity controlled by selector genes and intercellular signals. skuld and kohtalo encode homologs of TRAP240 and TRAP230, the two largest subunits of the Drosophila mediator complex; mutations in either gene cause identical phenotypes. We show here that both genes are required to establish normal cell affinity differences at the anterior-posterior and dorsal-ventral compartment boundaries of the wing disc. Mutant cells cross from the anterior to the posterior compartment, and can distort the dorsal-ventral boundary in either the dorsal or ventral direction. The Skuld and Kohtalo proteins physically interact in vivo and have synergistic effects when overexpressed, consistent with a skuld kohtalo double-mutant phenotype that is indistinguishable from either single mutant. We suggest that these two subunits do not participate in all of the activities of the mediator complex, but form a submodule that is required to regulate specific target genes, including those that control cell affinity.

Signaling by the secreted hedgehog, decapentaplegic and wingless proteins organizes the pattern of photoreceptor differentiation within the Drosophila eye imaginal disc; hedgehog and decapentaplegic are required for differentiation to initiate at the posterior margin and progress across the disc, while wingless prevents it from initiating at the lateral margins. Our analysis of these interactions has shown that initiation requires both the presence of decapentaplegic and the absence of wingless, which inhibits photoreceptor differentiation downstream of the reception of the decapentaplegic signal. However, wingless is unable to inhibit differentiation driven by activation of the epidermal growth factor receptor pathway. The effect of wingless is subject to regional variations in control, as the anterior margin of the disc is insensitive to wingless inhibition. The eyes absent and eyegone genes encode members of a group of nuclear proteins required to specify the fate of the eye imaginal disc. We show that both eyes absent and eyegone are required for normal activation of decapentaplegic expression at the posterior and lateral margins of the disc, and repression of wingless expression in presumptive retinal tissue. The requirement for eyegone can be alleviated by inhibition of the wingless signaling pathway, suggesting that eyegone promotes eye development primarily by repressing wingless. These results provide a link between the early specification and later differentiation of the eye disc.

The gene for rhodopsin, the primary light sensor of the visual system, is specifically expressed in the rod photoreceptor cells of the retina. We show here that in the rat, opsin RNA first accumulates to detectable levels at postnatal day 2 (PN2) and that nascent transcripts can be detected at PN1; this is the time when peak numbers of photoreceptor cells are generated by the final division of their neuroepithelial precursors. Accumulated opsin RNA then increases to reach the adult level, 0.06% of total retinal RNA, at about PN10. The transcription rate of the opsin gene increases to a similar extent over the same time course between PN3 and adulthood, suggesting that transcriptional activation is responsible for the increase in opsin expression. We used the antibody RET-P1 to show that rhodopsin protein is also detectable at PN2 and that the number of cells expressing the protein increases with time in a central-to-peripheral gradient in the retina. This increase in the number of differentiating photoreceptors in the tissue appears to account for much of the increase in opsin gene transcription and RNA accumulation. In situ hybridization to opsin RNA shows that it is restricted to the photoreceptor layer from the time it can first be detected, at PN7. Later in development, when RET-P1 staining shifts to the photoreceptor outer segments, opsin RNA becomes localized to the inner segments, suggesting that the distributions of opsin protein and RNA are related.