Kaoru Sugimura

Elimination of Oncogenic Neighbors by JNK-Mediated Engulfment in Drosophila

A newly emerged oncogenic cell in the epithelial population has to confront antitumor selective pressures in the host tissue. However, the mechanisms by which surrounding normal tissue exerts antitumor effects against oncogenically transformed cells are poorly understood. In Drosophila imaginal epithelia, clones of cells mutant for evolutionarily conserved tumor suppressor genes such as scrib or dlg lose their epithelial integrity and are eliminated from epithelia when surrounded by wild-type tissue. Here, we show that surrounding normal cells activate nonapoptotic JNK signaling in response to the emergence of oncogenic mutant cells. This JNK activation leads to upregulation of PVR, the Drosophila PDGF/VEGF receptor. Genetic and time-lapse imaging analyses reveal that PVR expression in surrounding cells activates the ELMO/Mbc-mediated phagocytic pathway, thereby eliminating oncogenic neighbors by engulfment. Our data indicate that JNK-mediated cell engulfment could be an evolutionarily conserved intrinsic tumor-suppression mechanism that eliminates premalignant cells from epithelia.

Shortstop Recruits EB1/APC1 and Promotes Microtubule Assembly at the Muscle-Tendon Junction

BACKGROUND Shot (previously named Kakapo), is a Drosophila Plakin family member containing both Actin binding and microtubule binding domains. In Drosophila, it is required for a wide range of processes, including axon extension, dendrite formation, axonal terminal arborization at the neuromuscular junction, tendon cell development, and adhesion of wing epithelium. RESULTS To address how Shot exerts its activity at the molecular level, we investigated the molecular interactions of Shot with candidate proteins in mature larval tendon cells. We show that Shot colocalizes with EB1/APC1 and with a compact microtubule array extending between the muscle-tendon junction and the cuticle. Shot forms a protein complex with EB1 via its C-terminal EF-hands and GAS2-containing domains. In tendon cells with reduced Shot activity, EB1/APC1 dissociate from the muscle-tendon junction, and the microtubule array elongates. The resulting tendon cell, although associated with the muscle and the cuticle ends, loses its stress resistance and elongates. CONCLUSIONS Our results suggest that Shot mediates tendon stress resistance by the organization of a compact microtubule network at the muscle-tendon junction. This is achieved by Shot association with the cytoplasmic faces of the basal hemiadherens junction and with the EB1/APC1 complex.

Bayesian inference of force dynamics during morphogenesis

During morphogenesis, cells push and pull each other to trigger precise deformations of a tissue to shape the body. Therefore, to understand the development of animal forms, it is essential to analyze how mechanical forces coordinate behaviors of individual cells that underlie tissue deformations. However, the lack of a direct and non-invasive force-measurement method has hampered our ability to identify the underlying physical principles required to regulate morphogenesis. In this study, by employing Bayesian statistics, we develop a novel inverse problem framework to estimate the pressure of each cell and the tension of each contact surface from the observed geometry of the cells. We confirmed that the true and estimated values of forces fit well in artificially generated data sets. Moreover, estimates of forces in Drosophila epithelial tissues are consistent with other readouts of forces obtained by indirect or invasive methods such as laser-induced destruction of cortical actin cables. Using the method, we clarify the developmental changes in the patterns of tensile force in the Drosophila dorsal thorax. In summary, the batch and noninvasive nature of the described force-estimation method will enable us to analyze the mechanical control of morphogenesis at an unprecedented quantitative level.

Development of Morphological Diversity of Dendrites in Drosophila by the BTB-Zinc Finger Protein Abrupt

Morphological diversity of dendrites contributes to specialized functions of individual neurons. In the present study, we examined the molecular basis that generates distinct morphological classes of Drosophila dendritic arborization (da) neurons. da neurons are classified into classes I to IV in order of increasing territory size and/or branching complexity. We found that Abrupt (Ab), a BTB-zinc finger protein, is expressed selectively in class I cells. Misexpression of ab in neurons of other classes directed them to take the appearance of cells with smaller and/or less elaborated arbors. Loss of ab functions in class I neurons resulted in malformation of their typical comb-like arbor patterns and generation of supernumerary branch terminals. Together with the results of monitoring dendritic dynamics of ab-misexpressing cells or ab mutant ones, all of the data suggested that Ab endows characteristics of dendritic morphogenesis of the class I neurons.

Selective expression of Knot/Collier, a transcriptional regulator of the EBF/Olf-1 family, endows the Drosophila sensory system with neuronal class-specific elaborated dendritic patterns.

Dendritic tree morphology is a hallmark of cellular diversity in the nervous system, and Drosophila dendritic arborization (da) neurons provide an excellent model system to study its molecular basis. The da neurons are classified into four classes I–IV in the order of increasing branching complexity. A transcriptional regulator of the early B‐cell factor (EBF)/olfactory 1 (Olf‐1) family, Knot (Kn)/Collier (Col) is expressed selectively in class IV neurons, which generate the most expansive and complicated dendritic trees in the four classes. Loss of kn function in class IV neurons greatly reduced the number of their dendritic branches. Conversely mis‐expression of kn in classes I and II produced supernumerary higher‐order branches, whereas class III‐specific short and straight terminal branches was hardly formed by kn mis‐expression. Neither kn loss of function nor mis‐expression were associated with dramatic alterations in the expression patterns of two other transcriptional regulators, Abrupt (Ab) and Cut (Ct), which play important roles in shaping dendritic trees with distinct class specificity from Kn. In contrast, Kn was necessary and sufficient to drive expression of a gene that encodes a class IV‐specific channel protein. Collectively, all of our results suggest that Kn exerts its cell‐autonomous function to control the formation, and possibly the function, of class IV‐like elaborated dendritic arbors.

Measuring forces and stresses in situ in living tissues

Development, homeostasis and regeneration of tissues result from a complex combination of genetics and mechanics, and progresses in the former have been quicker than in the latter. Measurements of in situ forces and stresses appear to be increasingly important to delineate the role of mechanics in development. We review here several emerging techniques: contact manipulation, manipulation using light, visual sensors, and non-mechanical observation techniques. We compare their fields of applications, their advantages and limitations, and their validations. These techniques complement measurements of deformations and of mechanical properties. We argue that such approaches could have a significant impact on our understanding of the development of living tissues in the near future. Summary: This Primer summarizes the current range of invasive and non-invasive techniques to measure forces in cells and tissues, and discusses their applications in developmental contexts.

Unified quantitative characterization of epithelial tissue development

Understanding the mechanisms regulating development requires a quantitative characterization of cell divisions, rearrangements, cell size and shape changes, and apoptoses. We developed a multiscale formalism that relates the characterizations of each cell process to tissue growth and morphogenesis. Having validated the formalism on computer simulations, we quantified separately all morphogenetic events in the Drosophila dorsal thorax and wing pupal epithelia to obtain comprehensive statistical maps linking cell and tissue scale dynamics. While globally cell shape changes, rearrangements and divisions all significantly participate in tissue morphogenesis, locally, their relative participations display major variations in space and time. By blocking division we analyzed the impact of division on rearrangements, cell shape changes and tissue morphogenesis. Finally, by combining the formalism with mechanical stress measurement, we evidenced unexpected interplays between patterns of tissue elongation, cell division and stress. Our formalism provides a novel and rigorous approach to uncover mechanisms governing tissue development. DOI: http://dx.doi.org/10.7554/eLife.08519.001

Filamin acts as a key regulator in epithelial defence against transformed cells

Recent studies have shown that certain types of transformed cells are extruded from an epithelial monolayer. However, it is not known whether and how neighbouring normal cells play an active role in this process. In this study, we demonstrate that filamin A and vimentin accumulate in normal cells specifically at the interface with Src- or RasV12-transformed cells. Knockdown of filamin A or vimentin in normal cells profoundly suppresses apical extrusion of the neighbouring transformed cells. In addition, we show in zebrafish embryos that filamin plays a positive role in the elimination of the transformed cells. Furthermore, the Rho/Rho kinase pathway regulates filamin accumulation and filamin acts upstream of vimentin in the apical extrusion. This is the first report demonstrating that normal epithelial cells recognize and actively eliminate neighbouring transformed cells and that filamin is a key mediator in the interaction between normal and transformed epithelial cells.

The mechanical anisotropy in a tissue promotes ordering in hexagonal cell packing.

Many epithelial tissues pack cells into a honeycomb pattern to support their structural and functional integrity. Developmental changes in cell packing geometry have been shown to be regulated by both mechanical and biochemical interactions between cells; however, it is largely unknown how molecular and cellular dynamics and tissue mechanics are orchestrated to realize the correct and robust development of hexagonal cell packing. Here, by combining mechanical and genetic perturbations along with live imaging and Bayesian force inference, we investigate how mechanical forces regulate cellular dynamics to attain a hexagonal cell configuration in the Drosophila pupal wing. We show that tissue stress is oriented towards the proximal-distal axis by extrinsic forces acting on the wing. Cells respond to tissue stretching and orient cell contact surfaces with the stretching direction of the tissue, thereby stabilizing the balance between the intrinsic cell junction tension and the extrinsic force at the cell-population level. Consequently, under topological constraints of the two-dimensional epithelial sheet, mismatches in the orientation of hexagonal arrays are suppressed, allowing more rapid relaxation to the hexagonal cell pattern. Thus, our results identify the mechanism through which the mechanical anisotropy in a tissue promotes ordering in cell packing geometry.

Nonautonomous Apoptosis Is Triggered by Local Cell Cycle Progression during Epithelial Replacement in Drosophila

ABSTRACT Tissue remodeling involves collective cell movement, and cell proliferation and apoptosis are observed in both development and disease. Apoptosis and proliferation are considered to be closely correlated, but little is known about their coordinated regulation in physiological tissue remodeling in vivo. The replacement of larval abdominal epidermis with adult epithelium in Drosophila pupae is a simple model of tissue remodeling. During this process, larval epidermal cells (LECs) undergo apoptosis and are replaced by histoblasts, which are adult precursor cells. By analyzing caspase activation at the single-cell level in living pupae, we found that caspase activation in LECs is induced at the LEC/histoblast boundary, which expands as the LECs die. Manipulating histoblast proliferation at the LEC/histoblast boundary, either genetically or by UV illumination, indicated that local interactions with proliferating histoblasts triggered caspase activation in the boundary LECs. Finally, by monitoring the spatiotemporal dynamics of the S/G2/M phase in histoblasts in vivo, we found that the transition from S/G2 phases is necessary to induce nonautonomous LEC apoptosis at the LEC/histoblast boundary. The replacement boundary, formed as caspase activation is regulated locally by cell-cell communication, may drive the dynamic orchestration of cell replacement during tissue remodeling.