John S. Poulton

PtdIns(3)P controls cytokinesis through KIF13A-mediated recruitment of FYVE-CENT to the midbody

Several subunits of the class III phosphatidylinositol-3-OH kinase (PI(3)K-III) complex are known as tumour suppressors. Here we uncover a function for this complex and its catalytic product phosphatidylinositol-3-phosphate (PtdIns(3)P) in cytokinesis. We show that PtdIns(3)P localizes to the midbody during cytokinesis and recruits a centrosomal protein, FYVE-CENT (ZFYVE26), and its binding partner TTC19, which in turn interacts with CHMP4B, an endosomal sorting complex required for transport (ESCRT)-III subunit implicated in the abscission step of cytokinesis. Translocation of FYVE-CENT and TTC19 from the centrosome to the midbody requires another FYVE-CENT-interacting protein, the microtubule motor KIF13A. Depletion of the VPS34 or Beclin 1 subunits of PI(3)K-III causes cytokinesis arrest and an increased number of binucleate and multinucleate cells, in a similar manner to the depletion of FYVE-CENT, KIF13A or TTC19. These results provide a mechanism for the translocation and docking of a cytokinesis regulatory machinery at the midbody.

Involvement of Lgl and Mahjong/VprBP in Cell Competition

Mahjong is a novel Lethal giant larvae-binding protein that plays a vital role in cell competition in both flies and mammals.

Perlecan and Dystroglycan act at the basal side of the Drosophila follicular epithelium to maintain epithelial organization

Dystroglycan (Dg) is a widely expressed extracellular matrix (ECM) receptor required for muscle viability, synaptogenesis, basementmembrane formation and epithelial development. As an integral component of the Dystrophin-associated glycoprotein complex, Dg plays a central role in linking the ECM and the cytoskeleton. Disruption of this linkage in skeletal muscle leads to various types of muscular dystrophies. In epithelial cells, reduced expression of Dg is associated with increased invasiveness of cancer cells. We have previously shown that Dg is required for epithelial cell polarity in Drosophila, but the mechanisms of this polarizing activity and upstream/downstream components are largely unknown. Using the Drosophila follicle-cell epithelium (FCE) as a model system, we show that the ECM molecule Perlecan (Pcan) is required for maintenance of epithelial-cell polarity. Follicle cells that lack Pcan develop polarity defects similar to those of Dg mutant cells. Furthermore, Dg depends on Pcan but not on Laminin A for its localization in the basal-cell membrane, and the two proteins bind in vitro. Interestingly, the Dg form that interacts with Pcan in the FCE lacks the mucin-like domain, which is thought to be essential for Dg ligand binding activity. Finally, we describe two examples of how Dg promotes the differentiation of the basal membrane domain: (1) by recruiting/anchoring the cytoplasmic protein Dystrophin; and (2) by excluding the transmembrane protein Neurexin. We suggest that the interaction of Pcan and Dg at the basal side of the epithelium promotes basal membrane differentiation and is required for maintenance of cell polarity in the FCE.

The hippo pathway promotes Notch signaling in regulation of cell differentiation, proliferation, and oocyte polarity.

Specification of the anterior-posterior axis in Drosophila oocytes requires proper communication between the germ-line cells and the somatically derived follicular epithelial cells. Multiple signaling pathways, including Notch, contribute to oocyte polarity formation by controlling the temporal and spatial pattern of follicle cell differentiation and proliferation. Here we show that the newly identified Hippo tumor-suppressor pathway plays a crucial role in the posterior follicle cells in the regulation of oocyte polarity. Disruption of the Hippo pathway, including major components Hippo, Salvador, and Warts, results in aberrant follicle-cell differentiation and proliferation and dramatic disruption of the oocyte anterior-posterior axis. These phenotypes are related to defective Notch signaling in follicle cells, because misexpression of a constitutively active form of Notch alleviates the oocyte polarity defects. We also find that follicle cells defective in Hippo signaling accumulate the Notch receptor and display defects in endocytosis markers. Our findings suggest that the interaction between Hippo and classic developmental pathways such as Notch is critical to spatial and temporal regulation of differentiation and proliferation and is essential for development of the body axes in Drosophila.

Deconstructing the ßcatenin destruction complex: mechanistic roles for the tumor suppressor APC in regulating Wnt signaling

APC is a key tumor suppressor and Wnt signaling regulator, but its mechanism of action remains mysterious. We combined parallel assays in Drosophila and cultured human colon cancer cell lines to test hypotheses regarding APC function and to develop novel hypotheses, using mutants altering its structure in specific ways.

Acentrosomal Drosophila Epithelial Cells Exhibit Abnormal Cell Division, Leading to Cell Death and Compensatory Proliferation

Mitotic spindles are critical for accurate chromosome segregation. Centrosomes, the primary microtubule nucleating centers of animal cells, play key roles in forming and orienting mitotic spindles. However, the survival of Drosophila without centrosomes suggested they are dispensable in somatic cells, challenging the canonical view. We used fly wing disc epithelia as a model to resolve these conflicting hypotheses, revealing that centrosomes play vital roles in spindle assembly, function, and orientation. Many acentrosomal cells exhibit prolonged spindle assembly, chromosome missegregation, DNA damage, misoriented divisions, and eventual apoptosis. We found that multiple mechanisms buffer the effects of centrosome loss, including alternative microtubule nucleation pathways and the spindle assembly checkpoint. Apoptosis of acentrosomal cells is mediated by JNK signaling, which also drives compensatory proliferation to maintain tissue integrity and viability. These data reveal the importance of centrosomes in fly epithelia and demonstrate the robust compensatory mechanisms at the cellular and organismal level.

Dystroglycan down-regulation links EGF receptor signaling and anterior–posterior polarity formation in the Drosophila oocyte

Anterior–posterior axis formation in the Drosophila oocyte requires activation of the EGF receptor (EGFR) pathway in the posterior follicle cells (PFC), where it also redirects them from the default anterior to the posterior cell fate. The relationship between EGFR activity in the PFC and oocyte polarity is unclear, because no EGFR-induced changes in the PFC have been observed that subsequently affect oocyte polarity. Here, we show that an extracellular matrix receptor, Dystroglycan, is down-regulated in the PFC by EGFR signaling, and this down-regulation is necessary for proper localization of posterior polarity determinants in the oocyte. Failure to down-regulate Dystroglycan disrupts apicobasal polarity in the PFC, which includes mislocalization of the extracellular matrix component Laminin. Our data indicate that Dystroglycan links EGFR-induced repression of the anterior follicle cell fate and anterior–posterior polarity formation in the oocyte.

Interphase centrosome organization by the PLP-Cnn scaffold is required for centrosome function

Cnn and PLP directly interact at two defined sites to coordinate the cell cycle–dependent rearrangement and scaffolding activity of the centrosome to permit normal centrosome organization, cell division, and embryonic viability.

Regulation of Wnt signaling by the tumor suppressor adenomatous polyposis coli does not require the ability to enter the nucleus or a particular cytoplasmic localization.

In this study, we test two current models for the function of the tumor suppressor adenomatous polyposis coli (APC). We find that APC can regulate Wnt signaling from diverse cytoplasmic locations, suggesting that its roles in the nucleus or in localizing the β-catenin destruction complex are not essential.

Regulation of Wnt signaling by the tumor suppressor APC does not require ability to enter the nucleus nor a particular cytoplasmic localization

In this study, we test two current models for the function of the tumor suppressor adenomatous polyposis coli (APC). We find that APC can regulate Wnt signaling from diverse cytoplasmic locations, suggesting that its roles in the nucleus or in localizing the β-catenin destruction complex are not essential.