Jérémie Rossy

Fluorescence imaging of a T cell

Endosomal networks in immunology

When a T cell in our immune system gets the right signal it leaps into action within seconds - taking on one of many different roles in fighting the infection.

Jérémie Rossy and his group are using single molecule imaging to examine how messages received at the cell surface are transmitted via the membrane and specialised compartments within the cell, such as endosomes.

“So far, the involvement of endosomes in transmitting messages within the cell has been overlooked by scientists,” says Jérémie.

He believes there’s no better way to understand how a cell responds to these messages than to be able to visualise the cell changing, and seeing where and when these changes take place.

The lab is currently working on three different projects

1) Is T cell signalling regulated by an activated endosomal network? The engagement of the T cell receptor (TCR) results in diverse cellular and immunological outcomes, from self-tolerance and killing of target cells to immunological memory. Consequently, the signals originating from the TCR and its co-receptors must be fed into and differentially processed by a complex network of signalling pathways. However, how TCR engagement results in various cellular responses remains poorly understood. The endocytic matrix represents a powerful cellular mechanism to shape the strength and outcome of signals generated at the plasma membrane. We and others have shown that in T cells too, effectors contained in intracellular vesicles participate in signalling beyond the plasma membrane (Williamson, Owen, Rossy et al., Nat Immunol, 2011). We believe that the reorganisation of the plasma membrane and intracellular compartments after engagement of the TCR serves to establish a specialised endosomal network, where the amplification, diversification and consolidation of TCR signalling take place. We have developed a strategy based on live-cell imaging, single molecule imaging and electron microscopy to uncover how endosomes contribute to T cell signalling.

2) Understanding the initiation of T cell activation. T lymphocytes participate in an immune response when they become activated through the TCR. However, even more than 25 years after the discovery of the TCR, the question of “How does T cell receptor signalling begin”? remains incompletely understood despite the identification of the major players and sequences of events. It is, for example, well established that TCR signalling is initiated when peptides bound to major histocompatibility complexes (pMHC) engage the receptor. It is also accepted that the first signalling event is the phosphorylation of the intracellular domains of TCR by the Src kinase Lck. However, for this event to occur, the kinase and substrate must be in close proximity to interact, and to initiate and sustain signalling; yet the underlying mechanisms for this molecular process are unknown. Using single molecule imaging, we have demonstrated that TCR activation triggers the clustering of Lck, and that this clustering is controlled by the conformation of the kinase, the active/open form inducing clustering and the inactive/closed form preventing it (Rossy et al, Nat. Immunol, 2013) The aim of this project is to extend these results and understand the dynamics of Lck clustering as well as the raison d'être of the Lck clusters.

3) What tells T cells when to stop? T cells circulate between the bloodstream and lymphoid organs. In the lymph node, T cells migrate at a staggering speed of 10-15 µm/min and constantly scan dendritic cells in search of cognate antigens. T cells can scan up to 5000 dendritic cells/h, which makes them the most highly migratory tissue cells in an adult body. But while a T cell reviews as many dendritic cells as possible, it must be able to slow down and eventually stop migrating when its TCR engages with a cognate peptide presented at the surface of a dendritic cell. Thus, the adaptive immune response relies on the ability of a few rapidly moving antigen-specific T cells to establish a stable interaction - the immunological synapse - with a limited number of appropriate dendritic cells.

TCR engagement can be regarded as the turning point of two cellular processes: it switches T cells from a motile to a stationary state and it converts ‘resting’ T cells into activated T cells, triggering TCR signalling and downstream activation responses. T cell migration in the lymph node generates extensive tensions and forces, which have a tremendous impact on cell membranes and signalling. To investigate the mechanistic link between migration-induced forces and T cell activation, we have established a multidisciplinary approach involving optical tweezers (collaboration with P. Reece, School of Physics, UNSW) and advanced imaging.

About Jérémie Rossy

After completing a master’s degree in Biochemistry at the University of Geneva, he focused his research on understanding immune cell signalling. During his PhD at the University of Bern, Jérémie used florescence microscopy to demonstrate that specialised domains are established and maintained in the plasma membrane of leukocytes, and that they control polarisation and migration of these cells. He moved to the University of New South Wales in 2009 to develop cutting-edge super-resolution microscopy techniques in order to gain novel insights into early stages of T lymphocyte activation.

Publications

More Information

Lab Members

  • Ewoud Compeer (postdoctoral fellow) - Membrane organising proteins as a link between the plasma membrane and endosomes
  • Natasha Kaushik (research assistant) - Dynamic of the kinase Lck within the immunological synapse