Visualising complex cell shapes and signalling pathways
Zebrafish larvae 24 hours after infection with fluorescent bacteria. Normal fish survive and eventually clear the bacteria (left), but if the interferon signalling pathway is compromised (right) the bacteria proliferate and the fish die
Two systems to study cell shape. Left: cross section of a 3-hour old embryo in which the ventral cells are beginning to invaginate; about 100 cells. Right: In the single, highly branched terminal tracheal cell the ER (blue) is involved in delivering membrane to the cells outer and inner (red) plasma membrane
The Leptin group studies the mechanisms and forces that determine cell shape in Drosophila and uses the zebrafish to analyse innate immune signalling.
In vivo imaging of innate immune responses
The innate immune system provides rapid defence against pathogens and also deals with non-pathogenic stresses. Macrophages and dendritic cells, two key players in this system, patrol the body and respond to stimuli from damaged cells via extra- and intracellular sensors. We aim to understand how such signals are recognised and how the appropriate subcellular and intercellular responses are triggered. We have discovered that one family of sensors, the cytoplasmic NOD-like receptors (NLRs), are particularly abundant in fish.
The zebrafish and medaka model systems allow in vivo observation of physiological processes. Specifically, we can watch pathogens and the cells that attack them. By genetic and chemical engineering we will generate in vivo fluorescent reporters for immune signalling events. These will be used to assay immune and stress responses in real time and at high spatial and temporal resolution as the cells of the fish encounter pathogens and stress signals.
Cell shape determination during development
The shape of a developing organism is generated by the activities of its constituent cells: growth and proliferation, movements, and shape changes. We are particularly interested in shape changes.
One study concerns an extremely complex single cell – the terminal cell of the Drosophila tracheal system. It is highly branched, and carries air to target tissues through an intracellular tube bounded by plasma membrane. During its rapid growth, the cell faces the task of synthesising large amounts of membrane and sorting it correctly to the outer and inner membrane domains. Extensive reorganisation of the secretory organelles precedes membrane growth. The cytoskeleton, small GTPases, and polarity determinants direct the process.
In another project, we try to understand how the forces generated by individual cells are integrated within the supracellular organisation of a tissue to give the tissue its final shape. We study the formation of the ventral furrow in the early embryo, which is well understood in regard to its genetics and cell biology. The cells that form the furrow are the major force generators driving invagination, but to allow furrow formation the neighbouring cells must respond, and they may contribute.
New genes we are currently discovering through a genetic screen for mRNAs localised in the branches of tracheal cells will be used for two purposes: a bioinformatic study of the signals that guide mRNAs to their specific subcellular localisation; and genetic and cell biological studies on how they contribute to branching and tube formation at that location. In vivo imaging with multicolour probes will be used to analyse the cellular mechanisms. To understand force integration across many cell populations, we will use quasi-simultaneous time-lapse imaging of multiple-angle views of the gastrulating embryo. We will measure the specific shape changes in all the cells of the embryo. We use genetic and mechanical manipulations to reveal the underlying control circuits. These studies are complemented by computational modelling.