UBC Theses and Dissertations

UBC Theses Logo

UBC Theses and Dissertations

The role of STAC proteins in regulation of calcium channels and skeletal muscle excitation-contraction coupling Rufenach-Barber, Britany

Abstract

Calcium is a ubiquitous signaling molecule that accompanies the cell from birth to death. Entry of Ca²⁺ into excitable cells is controlled by voltage-gated calcium channels (Caᵥs). STAC proteins have emerged in the last decade as novel regulators of these channels. They govern trafficking, current properties, and inactivation of L-type Caᵥs. Within this paradigm, the STAC3 isoform has a unique role: expressed solely in skeletal muscle, it has potential to be the long-sought link that allows for the unique mechanical coupling between Caᵥ1.1 and the ryanodine receptor (RyR1). Beyond muscle, other STAC isoforms are expressed in subsets of neurons where their physiological function is yet unknown. This thesis explores the roles of STAC proteins using structural biology and biophysical techniques to characterize interactions with a collection of proteins important for Ca²⁺ signaling. Variants in STAC3 lead to a rare yet debilitating neuromuscular disease known as STAC3 disorder. We use x-ray crystallography and isothermal titration calorimetry in conjunction with functional experiments to show these variants interrupt a critical interaction between STAC3 and Caᵥ1.1, thus impeding excitation-contraction coupling and Ca²⁺ release. Although the L-type current is dispensable for normal skeletal muscle function, it must be carefully controlled in other cell types (including neurons) to maintain precise Ca²⁺ homeostasis. STAC proteins affect this current by inhibiting the negative feedback mechanism of calcium-dependent inactivation (CDI). We present and characterize a novel interaction with another important regulatory protein – calmodulin (CaM). CaM is the resident Ca²⁺ sensor for L-type Caᵥs which triggers CDI. Given CaM’s ubiquitous expression in eukaryotic cells, this interaction may have widespread implications. Finally, shifting attention away from Caᵥs, we turn instead to the calcium channel on the other side of the triad: the ryanodine receptor. We use cryoEM and single-channel electrophysiology to explore whether STAC proteins may also fine-tune RyR1 function. Finding no evidence of a direct regulatory role of STAC on RyR1, we conclude the thesis with speculation on how STACs may cause disease, and how they may facilitate functional coupling between Caᵥ1.1 and RyR1 to enable that which powers our every movement and breath: skeletal muscle contraction.

Item Citations and Data

Rights

Attribution-NonCommercial-NoDerivatives 4.0 International