Leptin Group
Generation of complex cell shapes: membrane vesicles, polarity proteins and localised mRNAs
Below: a tracheal terminal cell expressing a fluorescent marker for the endoplasmic reticultum (blue). The lumen of the cell, which carries the oxygen for the surrounding (invisible) tissue, is seen in red (PhD thesis, Jayan Nair).
Cross section through a Drosophila embryo, in which the cells on the ventral side (bottom) have begun to change their shapes, creating an indentation that will eventually lead to the internalisation of these cells. The embryo is stained with antibodies against beta catenin (pink) and RhoGEF2 (blue). Image by Verena Kölsch.
Previous and current research
Many cells have highly polarised morphologies that serve specialised cell functions. We use the terminal cells of the Drosophila respiratory (tracheal) system and the epithelium of the gastrulating embryo to study the processes that determine cell shapes.
Terminal cells, like neurons, are extensively branched. The mechanisms involved in establishing their cellular architecture, especially the oxygen-carrying lumen, are poorly understood, as is also the case for related events in vertebrate tissues. Tracheal cell morphology depends on environmental signals that trigger events at sites distant from the cell body. We therefore assume that it depends on protein synthesis from locally stored RNA. We have developed an in vivo genetic screening method to search for such mRNAs. A pilot screen has identified genes with localised mRNAs that are required for the elaboration of the tracheal branches, and that had not been found by other methods. The analysis of these genes is beginning to reveal the mechanisms by which branches grow in response to local cues and generate the intracellular lumen to transport oxygen. These involve vesicle trafficking and fusion, definition of membrane domains and their polarity, and interactions of membranes with the cytoskeleton.
In the embryo, we have found that cell shape changes depend on localised G-protein dependent recruitment of cytoskeletal regulators, and require a dramatic re-distribution of adherens junctions by as yet unknown mechanisms. This redistribution occurs under the control of the transcription factor Snail, which controls epithelial-mesenchymal transitions in normal development and cancer, but the target genes through which Snail acts are largely unknown.
Future projects and goals
Our work on tracheal cells has two aims: the understanding of complex cell morphology and the search for signals involved in mRNA localisation. We will conduct a large-scale screen to identify all genes with localised mRNAs in tracheal cells, with a view to analysing their cell biological functions as well as their localisation signals. Results from testing genes from the pilot screen in other cell types have indicated that there must be cell type specific mRNA recognition and processing systems. To build a sequence database of mRNAs with tissue-specific polar distributions, the data obtained through screening tracheal cells in our own group will be enriched by input from collaborating laboratories who will use the insertion stocks we generate to screen other cell types. The combined datasets will be used for the bioinformatic identification of localisation signals.
To understand how the transcription factor Snail induces epithelial mesenchymal transitions, we will study sets of genes that were found in ChIP-on-chip experiments and by expression profiling to be direct targets of Snail in the mesoderm. In parallel, we will use laser micromanipulations and genetic and cell biological approaches to determine the mechanisms by which the adherens junctions are disassembled and reassembled.