Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating
<p>Intracellular Ca<sup>2+</sup>-release from the sarcoplasmic reticulum (SR) occurs through highly specialised Ca<sup>2+</sup>-release channels called ryanodine receptors (RyR) in a process known as excitation-contraction coupling (EC-coupling) in striated muscle.</...
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| Format: | Thesis |
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2015
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| _version_ | 1817932060695724032 |
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| author | O'Brien, F |
| author2 | Sitsapesan, R |
| author_facet | Sitsapesan, R O'Brien, F |
| author_sort | O'Brien, F |
| collection | OXFORD |
| description | <p>Intracellular Ca<sup>2+</sup>-release from the sarcoplasmic reticulum (SR) occurs through highly specialised Ca<sup>2+</sup>-release channels called ryanodine receptors (RyR) in a process known as excitation-contraction coupling (EC-coupling) in striated muscle.</p> <p>The RyR is the main ion-channel mediating Ca<sup>2+</sup>-release from the SR but the SR also contains many other ion-channels and proteins, with many of unknown physiological role. For example, two SR K<sup>+</sup> channels, TRIC-A and TRIC-B, have recently been identified and are thought to play a key role in supporting intracellular Ca<sup>2+</sup> movements across the SR. The TRIC double knockout (DKO) mouse dies in embryonic heart failure and the TRIC-B knockout (KO) dies in respiratory failure immediately after birth, highlighting their important roles in different tissue types. The TRIC-A KO mouse survives until adulthood, however, there are SR structural abnormalities and compromised Ca<sup>2+</sup> release.</p> <p>I have focused on characterising and comparing the single-channel properties of the native SR K<sup>+</sup> channels from WT and TRIC-A KO mouse skeletal muscle. I have investigated if the SR K<sup>+</sup> channels can gate in a cooperative manner when multiple SR K<sup>+</sup> channels are present in the bilayer. Since TRIC and RyR channels are both located in high abundance in the junctional SR regions, I have examined whether the SR K<sup>+</sup> and RyR channels also gate in a cooperative manner when both are present in a bilayer. In addition to voltage-regulation of native SR K<sup>+</sup> channels, I have characterised the pH sensitivity of the SR K<sup>+</sup> channels from WT and TRIC-A KO tissue. Finally, I investigated if the mouse isoform of TRIC-A could be purified using the Wheat germ cell-free system and incorporated into bilayers without loss of function. </p> <p>In summary, this thesis describes novel mechanisms of regulation of ion-channels that are involved in the process of Ca<sup>2+</sup>-release from the SR. By focusing on the biophysical properties of SR K<sup>+</sup> channels, I have highlighted the complexity of the ionic fluxes that are thought to be required to maintain normal EC-coupling in striated muscle.</p> |
| first_indexed | 2024-03-06T19:14:33Z |
| format | Thesis |
| id | oxford-uuid:17e51bc1-5b1f-4b52-8e46-98c888a91e00 |
| institution | University of Oxford |
| last_indexed | 2024-12-09T03:31:55Z |
| publishDate | 2015 |
| record_format | dspace |
| spelling | oxford-uuid:17e51bc1-5b1f-4b52-8e46-98c888a91e002024-12-01T15:17:03ZBiophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gatingThesishttp://purl.org/coar/resource_type/c_db06uuid:17e51bc1-5b1f-4b52-8e46-98c888a91e00ORA Deposit2015O'Brien, FSitsapesan, R<p>Intracellular Ca<sup>2+</sup>-release from the sarcoplasmic reticulum (SR) occurs through highly specialised Ca<sup>2+</sup>-release channels called ryanodine receptors (RyR) in a process known as excitation-contraction coupling (EC-coupling) in striated muscle.</p> <p>The RyR is the main ion-channel mediating Ca<sup>2+</sup>-release from the SR but the SR also contains many other ion-channels and proteins, with many of unknown physiological role. For example, two SR K<sup>+</sup> channels, TRIC-A and TRIC-B, have recently been identified and are thought to play a key role in supporting intracellular Ca<sup>2+</sup> movements across the SR. The TRIC double knockout (DKO) mouse dies in embryonic heart failure and the TRIC-B knockout (KO) dies in respiratory failure immediately after birth, highlighting their important roles in different tissue types. The TRIC-A KO mouse survives until adulthood, however, there are SR structural abnormalities and compromised Ca<sup>2+</sup> release.</p> <p>I have focused on characterising and comparing the single-channel properties of the native SR K<sup>+</sup> channels from WT and TRIC-A KO mouse skeletal muscle. I have investigated if the SR K<sup>+</sup> channels can gate in a cooperative manner when multiple SR K<sup>+</sup> channels are present in the bilayer. Since TRIC and RyR channels are both located in high abundance in the junctional SR regions, I have examined whether the SR K<sup>+</sup> and RyR channels also gate in a cooperative manner when both are present in a bilayer. In addition to voltage-regulation of native SR K<sup>+</sup> channels, I have characterised the pH sensitivity of the SR K<sup>+</sup> channels from WT and TRIC-A KO tissue. Finally, I investigated if the mouse isoform of TRIC-A could be purified using the Wheat germ cell-free system and incorporated into bilayers without loss of function. </p> <p>In summary, this thesis describes novel mechanisms of regulation of ion-channels that are involved in the process of Ca<sup>2+</sup>-release from the SR. By focusing on the biophysical properties of SR K<sup>+</sup> channels, I have highlighted the complexity of the ionic fluxes that are thought to be required to maintain normal EC-coupling in striated muscle.</p> |
| spellingShingle | O'Brien, F Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating |
| title | Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating |
| title_full | Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating |
| title_fullStr | Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating |
| title_full_unstemmed | Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating |
| title_short | Biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum K+ channel gating |
| title_sort | biophysical characterisation of the mechanisms regulating sarcoplasmic reticulum k channel gating |
| work_keys_str_mv | AT obrienf biophysicalcharacterisationofthemechanismsregulatingsarcoplasmicreticulumkchannelgating |