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Characterisation of human TASK-1 mutations in atrial fibrillation

Perfect, Walter Edward (2019) Characterisation of human TASK-1 mutations in atrial fibrillation. Master of Science by Research (MScRes) thesis, University of Kent,. (KAR id:80549)

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Abstract

Atrial fibrillation (AF) is the most common cardiac arrhythmia, affecting over two million Americans and this is set to triple by 2050, incurring an estimated U.S. treatment cost of $6.65 billion yearly. AF is a chronic condition associated with electrical changes within the heart that are related to atrial remodeling caused by variation in gene expression. The TWIK related acid sensitive potassium channel 1 (TASK-1, K2P3.1) is a protein with the ability to induce AF. The manifestation of TASK-1 induced AF has been categorized by three factors: incorrect protein trafficking, defective K2P3.1 channel assembly and TASK-1 electrophysiological dysfunction. However, the genomic variations that can induce these changes is not well documented, and so this project is aimed at characterising TASK-1 in human AF. This was conducted by the creation of mutant TASK-1 constructs pathogenic in AF. These included mutation of a transmembrane domain (V123L), and two Kozak mutations (-3A>U and -4G>A). These mutants were proposed to effect channel conformation and protein translation. For those reasons these mutations were picked for investigation, along with a C-terminally truncated protein, and a N-terminally mutated protein as to investigate the effects of phosphorylation sites and 14.3.3 binding in the trafficking of the protein. These constructs were individually or co-expressed on the membrane of tsA-201 cells; where they underwent electrophysiological analysis and confocal microscopy imaging. Overall TASK-1 mutants were shown, to present themselves in varied states. The C-terminus truncation and -3A mutations suggested an increased expression, by providing a significant increase in current compared to the wildtype. Whilst V123L proved to be potentially pathogenic, by demonstrating a significantly decreased current plus a significantly depolarized reversal potential. However, the mutational effects of V123L were rectified by heterodimerization with wildtype TASK-1, as would occur in native atrial tissue. Additionally, TASK-1 proved dominant in heterodimer formation with TASK-3, reducing the current amplitude close to that of TASK-1 homodimers, yet, the reversal potential was between the two reversal potential figures for homodimers of TASK-1 and TASK-3. Confocal imaging experiments showed that wildtype TASK-1 channels were expressed at the cell membrane. However, both the N-terminus and Kozak mutations imaged, proved even localization at the membrane, with no intracellular retention visible, suggesting that by use of another protocol, a quantifiable increase in expression maybe provided compared to wildtype.

Item Type: Thesis (Master of Science by Research (MScRes))
Thesis advisor: Mathie, Alistair
Uncontrolled keywords: Cardiac electrophysiology
Subjects: Q Science
Divisions: Divisions > Division of Natural Sciences > Medway School of Pharmacy
SWORD Depositor: System Moodle
Depositing User: System Moodle
Date Deposited: 19 Mar 2020 11:10 UTC
Last Modified: 01 Oct 2022 23:00 UTC
Resource URI: https://kar.kent.ac.uk/id/eprint/80549 (The current URI for this page, for reference purposes)

University of Kent Author Information

Perfect, Walter Edward.

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