Investigating the role of Zn²⁺ in regulating the function of intracellular Ca²⁺-release channels
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The tightly regulated openings of the cardiac ryanodine receptor (RyR2) help to ensure that intracellular Ca²⁺- release from the sarcoplasmic reticulum (SR) can only occur when heart contractions are required. Usually this process is self-regulatory, where Ca²⁺ both activates and inhibits release of further Ca²⁺ from the SR. In the progression of heart failure some of this control is lost and in rest periods Ca²⁺ can leak from the SR into the cytosol. Recent evidence has suggested that Zn²⁺- dyshomeostasis may contribute to SR Ca²⁺- leak but the underlying mechanism is unclear. Using single channel electrophysiological studies in combination with live cell imaging of HEK 293 and fibroblasts, this study reveals that Zn²⁺, along with Ca²⁺ and the inhibitor Mg²⁺, plays a physiological role in the grading of Ca²⁺- release via RyR2. Importantly the data reveal that pathophysiological concentrations of Zn²⁺ (> 100pM) within the cytosol remove the requirement of Ca²⁺ to activate RyR2, resulting in irregular channel activity even in the presence of Mg²⁺. This increase in channel open probability due to Zn²⁺ is known to be associated with increased Ca²⁺- release events such as Ca²⁺ sparks suggesting that Zn²⁺ is a regulator of the SR Ca²⁺-leak current. A potential source of releasable Zn²⁺, which could modulate RyR2 activity in cardiomyocytes, are the acidic organelles (endosomes and lysosomes). This study provides key evidence that the two pore channels (TPCs), which are expressed on the surface of these organelles, are candidate channels for ligand-gated release of Zn²⁺. Importantly this research demonstrates that dysregulated Zn²⁺ homeostasis, resulting in elevated Zn²⁺ within the lysosome, has severe consequences upon cellular Ca²⁺- release from fibroblasts, which is primarily the result of Zn²⁺ acting as a pore blocker of TPC2. Together these data reveal a key role of Zn²⁺ as a second messenger which can regulate intracellular Ca²⁺- release in both health and disease.
Thesis, PhD Doctor of Philosophy
Embargo Date: 2023-09-24
Embargo Reason: Thesis restricted in accordance with University regulations. Print and electronic copy restricted until 24th September 2023
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