Volume-regulated ion channels in T lymphocytes

Cells need to regulate their volume in response to changes in osmotic stress for their survival. For example, cells shrinkage can cause molecular crowding and ion imbalance, which affects the movement of molecules and vesicles within the cell. Fluctuations in the concentration of ions inside the cell can also impact cell signaling by changing the gradients of ions across the cell membrane. To maintain the right cell volume, cells use a network of ion channels and transporters (ICTs) which help with water movement and allow cells to decrease their volume when exposed to hypotonic conditions in a process called regulatory volume decrease (RVD), as well as increase their volume when exposed to hypertonic conditions. In T lymphocytes, these volume-activated ICTs have been found in developing cells in the thymus and mature cells in peripheral lymphoid organs. However, the mechanisms of activation and the physiological role of these channels in T cells are unclear since these cells are rarely exposed to hypotonic conditions within our body. Therefore, further studies are necessary to understand how volume-activated ICTs impact immune cell function.

Novel functions of volume-activated ion channels in immune cell signaling

Volume-regulated anion channels (VRACs) are hexameric complexes composed by members of the LRRC8 family. LRRC8A is the essential component of VRACs, but it requires to assemble with other LRRC8 paralogs (i.e. LRRC8B-E) to form functional channels. VRACs can be activated by a decrease in the cytosolic ionic strength and allow the transport of various substrates such as halides ions, metabolites, second messengers, anticancer drugs, and antibiotics. However, it is still unclear how VRACs transport large substrates in normal conditions. Exploring this aspect of VRACs could reveal additional functions beyond regulating cell volume. LRRC8 channels in lymphocytes are believed to play a role in their development and function, but further research is needed to understand the mechanisms of how VRACs regulate immune cell signaling and function.

Our previous work identified the VRAC component LRRC8C as a novel cyclic dinucleotide (CDN) transporter in T cells (PMID: 35105987). CDNs are second messengers that play important signaling functions in prokaryotic and eukaryotic cells, and many years ago they were shown to induce T cell apoptosis through an unidentified mechanism.  Our findings revealed that: i) T cells produce the CDN 2’3’ cyclic GMP-AMP (cGAMP) upon TCR stimulation, ii) T cells use LRRC8C to influx cGAMP and activate the CDN sensor Stimulator of Interferon Genes (STING), iii) activation of STING in T cells leads to the accumulation of p53 which in turn induces cell cycle arrest and apoptosis, and iv) activation of the STING-p53 signaling in T cells contributes to the modulation of T cell-mediated responses in mouse models of influenza A virus infection and experimental autoimmune encephalomyelitis. We proposed that cGAMP uptake through LRRC8C and STING-p53 signaling is a novel inhibitory signaling pathway in T cells and adaptive immunity.

Our research team is currently expanding our studies on LRRC8 channels in T cells and other immune cells to uncover their role in immune cell signaling and function. To achieve this goal, we are collaborating with multiple research teams and using cross-disciplinary approaches including biochemistry, electrophysiology, detailed structure-function studies, molecular immunology, systems biology and genetic mouse models.

Discovery and characterization of novel ion channel regulators of immune cell function

It is estimated that membrane proteins represent about 27% of all human proteome, and they are targets for more than half of FDA approved drugs. However, ~41% of these proteins have not been yet classified and the function of many of them is still unknown. In fact, ion channels which constitute a small fraction (~15%) of all membrane proteins are successfully used as drug targets in neurological and cardiovascular disorders. To discover new ICT regulators of immune cell function, we are expanding our previous RNAi screens in primary T cells (PMID: 35440113) by using CRISPR screens to interrogate the entire membrane proteome (ICTs, membrane receptors and enzymes, and adhesion molecules) to identify novel potential modulators of immune cell function.