A biosensor for the activity of the "sheddase" TACE (ADAM17) reveals novel and cell type-specific mechanisms of TACE activation.

Diverse environmental conditions stimulate protein "shedding" from the cell surface through proteolytic cleavage. The protease TACE [tumor necrosis factor-α (TNFα)--converting enzyme, encoded by ADAM17] mediates protein shedding, thereby regulating the maturation and release of various extracellular substrates, such as growth factors and cytokines, that induce diverse cellular responses. We developed a FRET (fluorescence resonance energy transfer)-based biosensor called TSen that quantitatively reports the kinetics of TACE activity in live cells. In combination with chemical biology approaches, we used TSen to probe the dependence of TACE activation on the induction of the kinases p38 and ERK (extracellular signal-regulated kinase) in various epithelial cell lines. Using TSen, we found that disruption of the actin cytoskeleton in keratinocytes induced rapid and robust TSen cleavage and the accumulation of TACE at the plasma membrane. Cytoskeletal disruption also increased the cleavage of endogenous TACE substrates, including transforming growth factor-α. Thus, TSen is a useful tool for unraveling the mechanisms underlying the spatiotemporal activation of TACE in live cells.