Quantifying single cell lipid signaling kinetics after photo-stimulation

Studying the role of molecularly distinct lipid species in cell signaling remains challenging due to a scarcity of methods for performing quantitative lipid biochemistry in living cells. We have recently used lipid uncaging to quantify lipid-protein affinities and rates of lipid transbilayer movement and turnover in the diacylglycerol signaling pathway using population average time series data. So far, this approach does not allow to account for the cell-to-cell variability of cellular signaling responses. We here report a framework that allows to uniquely identify model parameters such diacylglycerol-protein affinities and transbilayer movement rates at the single cell level for a broad variety of structurally different diacylglycerol species. We find that lipid unsaturation degree and longer side chains generally correlate with faster lipid transbilayer movement and turnover and higher lipid-protein affinities. In summary, our work demonstrates how rate parameters and lipid-protein affinities can be quantified from single cell signaling trajectories with sufficient sensitivity to resolve the subtle kinetic differences caused by the chemical diversity of cellular signaling lipid pools.