Research

Control of the dynein-dependent transport along microtubules in cell formation

Epithelial cells form the basic structure in many organs of multicellular animals. After fertilization of a Drosophila egg, mitotic cleavage divisions lead to a syncytium in which many cell nuclei share a common a cytoplasm. In a process called cellularization, the syncytium becomes a single-layer epithelium of approx. 6000 cells (Fig. 1). Our work shows that a biomedically highly relevant protein kinase of the family of microtubule-concentrated serine / threonine (MAST) kinases plays an essential role in the formation of cells in cellularization. Our research found that the Drosophila MAST kinase, Drop out, is responsible for the phosphorylation of the microtubule motor Dynein leading to the working hypothesis that Drop out is a central signaling molecule linking changes in gene regulation to the transition of the syncytium into the epithelial stage of the early embryo.

Figure 1: Epitheliogenesis in the Drosophila embryo. The nuclei (blue) are localised at the cell cortex. Different membrane proteins (green-Neurotactin, red-Armadillo) are incorporated into the growing plasma membranes. This process results in the generation of a polarised epithelial monolayer within a period of less than 1 hour under laboratory conditions.

Intercellular communication during morphogenesis

Fibroblast-Growth factors (FGF) are secreted cell signaling molecules, which are essential for the communication of cells in multicellular organisms including humans. FGFs control tissue and organ homeostasis by regulating the proliferation, migration, and survival of cells. Abnormal FGF signaling results in severe diseases such as cancer, cystic fibrosis, dysplasia. The complexity of FGF molecules and their receptors in vertebrates hamper their functional analysis in higher organisms. Our work has discovered an FGF signaling system in Drosophila and it’s role controlling cell behaviors including epithelial-mesenchymal transition and collective cell migration during the development of the mesoderm (Fig.2).

Figure 2: Epithelial-Mesenchymal Transition (EMT) in the mesoderm. The mesoderm cells (green/yellow) udnergos a transition from polarised epithelial cells into mesenchymal (migrating) cells. With this transition goes along a dramatic reorganisation of the epithelial structure of the mesoderm cells that is dependent on FGF signaling.

Development of high resolution Light Sheet Microscopy (SPIM)

Biological processes are dynamic and take place on a subcellular, cellular and supracellular level. The ideal dynamic microscopic imaging should be able to take into account the different spatial and temporal requirements of biologically safe systems. Light sheet microscopy (single plane illumination microscopy SPIM) is ideally suited for the overall imaging of living samples with high temporal and spatial resolution. In SPIM, the light scattering that occurs during imaging of biological samples leads to limitations in terms of resolution and field of view of the imaging. To remedy this deficiency, the illuminating beam can be modified into a very fine sheet of light over a large field of view. In addition, the technology of Multi-View-SPIM (M-SPIM) can be used to record the object from different positions by using several lenses in combination with mechanical rotation. In our working group we have developed a new type of multiview tiling SPIM (MT-SPIM) that combines the M-SPIM technology with a 'tiling light sheet'. The MT-SPIM offers high-resolution, robust and rotation-free imaging of living samples. We were able to show that the MT-SPIM improves the axial resolution compared to the conventional M-SPIM by a factor of two and that this improvement in the axial resolution improves the automated segmentation of subcellular structures.

Figure 3: Imaging sample chamber of the Multiview Tiling Single Plane illumination microscope (MT-SPIM). The two illumination lenses send thin tiled light sheets (blue arrows) onto the sample from opposite directions. These light sheets are scanned through the sampe by moving the objective lenses. The signal is detected at an angle of 90° by two detection lenses (green arrows) to obtain two opposite views from the sample.