Robust T-cell signaling triggered on soft polydimethylsiloxane-supported lipid-bilayers

The T-cell receptor (TCR) is thought to be triggered either by mechano-transduction or local tyrosine phosphatase exclusion at cell-cell contacts. However, the effects of the mechanical properties of activating surfaces have only been tested for late-stage T-cell activation, and phosphatase segregation has mostly been studied on glass-supported lipid bilayers that favor imaging but are orders-of-magnitude stiffer than typical cells. We developed a method for attaching lipid bilayers to polydimethylsiloxane polymer supports, producing "soft bilayers" with physiological levels of mechanical resistance (Youngs modulus of 4 kPa). Comparisons of T-cell behavior on soft and glass-supported bilayers revealed that early calcium signaling is unaffected by substrate rigidity, implying that early steps in TCR triggering are not mechanosensitive. Robust phosphatase exclusion was observed on the soft bilayers, however, suggesting it likely occurs at cell-cell contacts. This work sets the stage for an imaging-based exploration of receptor signaling under conditions closely mimicking physiological cell-cell contact.