Ionic protein–lipid interactions are critical for the structure and function of membrane receptors, ion channels, integrins and many other proteins,  However, the regulatory mechanism of these interactions is largely unknown. Here we show that Ca2+ can bind directly to anionic phospholipids and thus modulate membrane protein function. The activation of T-cell antigen receptor–CD3 complex (TCR), a key membrane receptor for adaptive immunity, is regulated by ionic interactions between positively charged CD3 / cytoplasmic domains (CD3CD) and negatively charged phospholipids in the plasma membrane. Crucial tyrosines are buried in the membrane and are largely protected from phosphorylation in resting T cells. It is not clear how CD3CD dissociates from the membrane in antigen-stimulated T cells. The antigen engagement of even a single TCR triggers a Ca2+ influx and TCR-proximal Ca2+ concentration is higher than the average cytosolic Ca2+ concentration. Our biochemical, live-cell fluorescence resonance energy transfer and NMR experiments showed that an increase in Ca2+ concentration induced the dissociation of CD3CD from the membrane and the solvent exposure of tyrosine residues. As a consequence, CD3 tyrosine phosphorylation was significantly enhanced by Ca2+ influx. Moreover, when compared with wild-type cells, Ca2+ channel-deficient T cells had substantially lower levels of CD3 phosphorylation after stimulation. The effect of Ca2+ on facilitating CD3 phosphorylation is primarily due to the charge of this ion, as demonstrated by the fact that replacing Ca2+ with the non-physiological ion Sr2+ resulted in the same feedback effect. Finally, 31P NMR spectroscopy showed that Ca2+ bound to the phosphate group in anionic phospholipids at physiological concentrations, thus neutralizing the negative charge of phospholipids. Rather than initiating CD3 phosphorylation, this regulatory pathway of Ca2+ has a positive feedback effect on amplifying and sustaining CD3 phosphorylation and should enhance T-cell sensitivity to foreign antigens. Our study thus provides a new regulatory mechanism of Ca2+ to T-cell activation involving direct lipid manipulation.

 

Related Links:http://www.nature.com/nature/journal/v493/n7430/full/nature11699.html