Identification of the Amino Terminus of Neuronal Ca2+Channel α1 Subunits α1B and α1E as an Essential Determinant of G-Protein Modulation

• DOI: 10.1523/jneurosci.18-13-04815.1998
• Publication Date: 2018-04-03T00:11:44Z
• Authors: Karen M. Page; Carles Cantı́; Gary J. Stephens; Nicholas S. Berrow; Annette C. Dolphin
• Keywords: 0303 health sciences; 03 medical and health sciences

We have examined the basis for G-protein modulation of the neuronal voltage-dependent calcium channels (VDCCs) α1E and α1B. A novel PCR product of α1E was isolated from rat brain. This contained an extended 5′ DNA sequence and was subcloned onto the previously cloned isoform rbEII, giving rise to α1Elongwhose N terminus was extended by 50 amino acids. VDCC α1 subunit constructs were co-expressed with the accessory α2-δ and β2a subunits inXenopusoocytes and mammalian (COS-7) cells. The α1Elongshowed biophysical properties similar to those of rbEII; however, when G-protein modulation of expressed α1 subunits was induced by activation of co-expressed dopamine (D2) receptors with quinpirole (100 nm) in oocytes, or by co-transfection of Gβ1γ2 subunits in COS-7 cells, α1Elong, unlike α1E(rbEII), was found to be G-protein-modulated, in terms of both a slowing of activation kinetics and a reduction in current amplitude. However, α1Elongshowed less modulation than α1B, and substitution of the α1E1–50with the corresponding region of α1B1–55produced a chimera α1bEEEE, with G-protein modulation intermediate between α1Elongand α1B. Furthermore, deletion of the N-terminal 1–55 sequence from α1B produced α1BΔN1–55, which could not be modulated, thus identifying the N-terminal domain as essential for G-protein modulation. Taken together with previous studies, these results indicate that the intracellular N terminus of α1E1–50and α1B1–55is likely to contribute to a multicomponent site, together with the intracellular I–II loop and/or the C-terminal tail, which are involved in Gβγ binding and/or in subsequent modulation of channel gating.