From regulation to pathogenesis: employing fluorescent tools to study calcium

McCombs, Janet Elaine

Graduate Thesis Or Dissertation

From regulation to pathogenesis: employing fluorescent tools to study calcium Public Deposited

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Citeable URL: https://scholar.colorado.edu/concern/graduate_thesis_or_dissertations/tx31qh82b

Abstract

Ca²⁺ is a ubiquitous and highly regulated signaling second messenger in cells, involved in a number of processes and pathways. Due to the complexity of Ca²⁺ signaling and its organization within a cell, it is inherently difficult to study. A number of fluorescent tools have been developed throughout the years in order to overcome these complications. Such probes allow for imaging of rapid Ca²⁺ transients in real-time at the single cell level, enabling the separation of the spatio-temporal aspects of Ca²⁺ signals. As Ca²⁺ is essential to cellular regulation and function, it is unsurprising Ca²⁺ + homeostasis is often altered in disease. In the studies herein, we examined Ca²⁺ as it pertains to pathogenesis in order to further current understandings about how its dysregulation leads to disease progression. By employing a specifically targeted Ca²⁺ sensor, we monitored changes in intracellular store levels and release caused by mutations in the Alzheimer’s disease related protein presenilin-1. Our results suggest mutations have differential effects on cellular Ca²⁺ homeostasis and stress the importance of directly investigating compartmentalized Ca²⁺ as opposed to inferring intracellular levels based on release into the cytosol. In addition, Ca²⁺ transients caused by bacterial infection of host cells were studied to determine molecular mechanisms involved in Salmonella-induced Ca²⁺ fluxes, providing insights into how bacteria modulate Ca²⁺ pathways to facilitate their survival. Finally, in order to characterize Ca²⁺ regulation in mitochondria, we used a mitochondrial targeted Ca²⁺ sensor to determine the involvement of a newly identified protein in Ca²⁺ uptake into the organelle. The use of fluorescent tools to study Ca²⁺ sheds light on effects of disease on Ca²⁺ regulation and homeostasis, providing mechanistic details that may lead to therapeutics.

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