Drosophila Apc2 Is a Cytoskeletally-Associated Protein That Regulates Wingless Signaling in the Embryonic Epidermis

The tumor suppressor adenomatous polyposis coli (APC) negatively regulates Wingless (Wg)/Wnt signal transduction by helping target the Wnt effector β-catenin or its Drosophila homologue Armadillo (Arm) for destruction. In cultured mammalian cells, APC localizes to the cell cortex near the ends of microtubules. Drosophila APC (dAPC) negatively regulates Arm signaling, but only in a limited set of tissues. We describe a second fly APC, dAPC2, which binds Arm and is expressed in a broad spectrum of tissues. dAPC2's subcellular localization revealed colocalization with actin in many but not all cellular contexts, and also suggested a possible interaction with astral microtubules. For example, dAPC2 has a striking asymmetric distribution in neuroblasts, and dAPC2 colocalizes with assembling actin filaments at the base of developing larval denticles. We identified a dAPC2 mutation, revealing that dAPC2 is a negative regulator of Wg signaling in the embryonic epidermis. This allele acts genetically downstream of wg, and upstream of arm, dTCF, and, surprisingly, dishevelled. We discuss the implications of our results for Wg signaling, and suggest a role for dAPC2 as a mediator of Wg effects on the cytoskeleton. We also speculate on more general roles that APCs may play in cytoskeletal dynamics

Non-equivalent roles of Drosophila Frizzled and Dfrizzled2 in embryonic Wingless signal transduction

AbstractThe highly conserved Wnt family of growth factors is essential for generating embryonic pattern in many animal species [1]. In the fruit fly Drosophila, most Wnt-mediated patterning is performed by a single family member, Wingless (Wg), acting through its receptors Frizzled (Fz) and DFrizzled2 (Dfz2). In the ventral embryonic epidermis, Wg signaling generates two different cell-fate decisions: the production of diverse denticle types and the specification of naked cuticle separating the denticle belts. Mutant alleles of wg disrupt these cellular decisions separately [2], suggesting that some aspect of ligand–receptor affinity influences cell-fate decisions, or that different receptor complexes mediate the distinct cellular responses. Here, we report that overexpression of Dfz2, but not Fz, rescues the mutant phenotype of wgPE2, an allele that produces denticle diversity but no naked cuticle. Fz was able to substitute for Dfz2 only under conditions where the Wg ligand was present in excess. The wgPE2 mutant phenotype was also sensitive to the dosage of glycosaminoglycans, suggesting that the mutant ligand is excluded from the receptor complex when proteoglycans are present. We conclude that wild-type Wg signaling requires efficient interaction between ligand and the Dfz2–proteoglycan receptor complex to promote the naked cuticle cell fate

Tum/RacGAP50C provides a critical link between anaphase microtubules and the assembly of the contractile ring in Drosophila melanogaster

A central question in understanding cytokinesis is how the cleavage plane is positioned. Although the positioning signal is likely to be transmitted via the anaphase microtubule array to the cell cortex, exactly how the microtubule array determines the site of contractile ring formation remains unresolved. By analysing tum/RacGAP50C mutant Drosophila embryos we show that cells lacking Tum do not form furrows and fail to localise the key cytokinetic components Pebble (a RhoGEF), Aurora B kinase, Diaphanous, Pav-KLP and Anillin. The GAP activity of Tum is required for cytokinesis: in its absence cytokinesis fails early even though Tum is present on microtubules at the cell equator where the furrow should form. Disruption of the Pebble-interacting domain leaves Tum localised to the cell equator on cortically associated microtubules, again with no evidence of furrowing. These data support a model in which Tum/RacGAP, via its interaction with Pbl, provides a critical link between the anaphase microtubule spindle and cytokinetic furrow formation in Drosophila cells

Testing hypotheses for the functions of APC family proteins using null and truncation alleles in Drosophila.

Adenomatous polyposis coli (APC) is mutated in colon cancers. During normal development, APC proteins are essential negative regulators of Wnt signaling and have cytoskeletal functions. Many functions have been proposed for APC proteins, but these have often rested on dominant-negative or partial loss-of-function approaches. Thus, despite intense interest in APC, significant questions remain about its full range of cellular functions and about how mutations in the gene affect these. We isolated six new alleles of Drosophila APC2. Two resemble the truncation alleles found in human tumors and one is a protein null. We generated ovaries and embryos null for both APC2 and APC1, and assessed the consequences of total loss of APC function, allowing us to test several previous hypotheses. Surprisingly, although complete loss of APC1 and APC2 resulted in strong activation of Wingless signaling, it did not substantially alter cell viability, cadherin-based adhesion, spindle morphology, orientation or selection of division plane, as predicted from previous studies. We also tested the hypothesis that truncated APC proteins found in tumors are dominant negative. Two mutant proteins have dominant effects on cytoskeletal regulation, affecting Wnt-independent nuclear retention in syncytial embryos. However, they do not have dominant-negative effects on Wnt signaling.</p