Abstract
L-type calcium channels (LTCCs) are voltage-gated calcium channels made up of three subunits, the α1 pore-forming subunit and the β and α2δ auxiliary subunits. LTCC’s have a wide range of cellular functions, including activation of the calcium-dependent components of the medium and slow afterhyperpolarisations (AHPs) in hippocampal neurons. AHPs are important for maintaining normal neuronal firing patterns. Tau is a microtubule associated protein (MAP) with six isoforms which have either three or four microtubule binding repeats (3R and 4R), and zero, one or two amino terminal inserts (0-2N). Preliminary evidence shows that the ratio of 3R:4R tau is imbalanced in Alzheimer’s Disease (AD) and other dementias and causes an increase in number of LTCCs at the cell membrane of hippocampal neurons.Previous research has shown that 4R0N-tau and 4R2N-tau, but not 4R1N-tau, augments L-type calcium currents in tsA201 cells, 4R0N-tau having the strongest effect. This interaction only occurs in LTCCs comprised of the α1C subunit (Cav1.2) and Cavβ3 subunit isoforms and indicates that there is a direct interaction between 4R0N-tau and the Cavβ3 subunit. The aim of this research is to determine the site of interaction within the CaVβ3 subunit that mediates the proposed direct interaction between 4R0N-tau and CaVβ3.
Electrophysiological experiments using tSA201 cell lines exogenously expressing CaV1.2, β3, and α2δ1 elicited L-type calcium current that was augmented when co-expressed with 4R0N-tau. Further electrophysiological experiments were conducted to find a point mutation which removed the augmentation of L-type calcium currents by 4R0N-tau. The W102F mutation of Cavβ3 successfully achieved this aim without changing individual channel activity, assessed by calculating V0.5 and decay constant.
This research shows that mutation of a single tryptophan to phenylalanine, W102F, abolishes the ability of 4R0N-tau to augment L-type current. Increased concentrations of 4R0N-tau and upregulation of L type current has been reported in AD brains, and in aged animals experiencing cognitive decline, but not in aged animals that didn’t experience cognitive decline. Therefore, the identification of the specific residue where the interaction between 4R0N-tau and Cavβ3 helps our understanding of AD pathomechanism and offers a potential therapeutic target for prevention.
| Date of Award | 1 Oct 2024 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | David N Sheppard (Supervisor) |