ADAM17 mediates proteolytic maturation of voltage-gated calcium channel auxiliary α₂δ subunits, and enables calcium current enhancement

The auxiliary alpha(2)delta subunits of voltage-gated calcium (Ca-V) channels are key to augmenting expression and function of Ca(V)1 and Ca(V)2 channels, and are also important drug targets in several therapeutic areas, including neuropathic pain. The alpha(2)delta proteins are translated as pre-proteins encoding both alpha(2) and delta, and post-translationally proteolysed into alpha(2) and delta subunits, which remain associated as a complex. In this study we have identified ADAM17 as a key protease involved in proteolytic processing of pro-alpha(2)delta-1 and alpha(2)delta-3 subunits. We provide three lines of evidence: firstly, proteolytic cleavage is inhibited by chemical inhibitors of particular metalloproteases, including ADAM17. Secondly, proteolytic cleavage of both alpha(2)delta-1 and alpha(2)delta-3 is markedly reduced in cell lines by knockout of ADAM17 but not ADAM10. Thirdly, proteolytic cleavage is reduced by the N-terminal active domain of TIMP-3 (N-TIMP-3), which selectively inhibits ADAM17. We have found previously that proteolytic cleavage into mature alpha(2)delta is essential for the enhancement of Ca-V function, and in agreement, knockout of ADAM17 inhibited the ability of alpha(2)delta-1 to enhance both Ca(V)2.2 and Ca(V)1.2 calcium currents. Finally, our data also indicate that the main site of proteolytic cleavage of alpha(2)delta-1 is the Golgi apparatus, although cleavage may also occur at the plasma membrane. Thus, our study identifies ADAM17 as a key protease required for proteolytic maturation of alpha(2)delta-1 and alpha(2)delta-3, and thus a potential drug target in neuropathic pain.