The HOOK region of voltage-gated Ca(2+) channel β subunits senses and transmits PIP(2) signals to the gate

The β subunit of voltage-gated Ca(2+) (Ca(V)) channels plays an important role in regulating gating of the α1 pore-forming subunit and its regulation by phosphatidylinositol 4,5-bisphosphate (PIP(2)). Subcellular localization of the Ca(V) β subunit is critical for this effect; N-terminal–dependent membrane targeting of the β subunit slows inactivation and decreases PIP(2) sensitivity. Here, we provide evidence that the HOOK region of the β subunit plays an important role in the regulation of Ca(V) biophysics. Based on amino acid composition, we broadly divide the HOOK region into three domains: S (polyserine), A (polyacidic), and B (polybasic). We show that a β subunit containing only its A domain in the HOOK region increases inactivation kinetics and channel inhibition by PIP(2) depletion, whereas a β subunit with only a B domain decreases these responses. When both the A and B domains are deleted, or when the entire HOOK region is deleted, the responses are elevated. Using a peptide-to-liposome binding assay and confocal microscopy, we find that the B domain of the HOOK region directly interacts with anionic phospholipids via polybasic and two hydrophobic Phe residues. The β2c-short subunit, which lacks an A domain and contains fewer basic amino acids and no Phe residues in the B domain, neither associates with phospholipids nor affects channel gating dynamically. Together, our data suggest that the flexible HOOK region of the β subunit acts as an important regulator of Ca(V) channel gating via dynamic electrostatic and hydrophobic interaction with the plasma membrane.