The proteostasis interactomes of trafficking-deficient variants of the voltage-gated potassium channel KV11.1 associated with long QT syndrome.

The voltage-gated potassium ion channel KV11.1 plays a critical role in cardiac repolarization. Genetic variants that render Kv11.1 dysfunctional cause long QT syndrome (LQTS), which is associated with fatal arrhythmias. Approximately 90% of LQTS-associated variants cause intracellular protein transport (trafficking) dysfunction, which pharmacological chaperones like E-4031 can rescue. Protein folding and trafficking decisions are regulated by chaperones, protein quality control factors, and trafficking machinery comprising the cellular proteostasis network. Here, we test whether trafficking dysfunction is associated with alterations in the proteostasis network of pathogenic Kv11.1 variants and whether pharmacological chaperones can normalize the proteostasis network of responsive variants. We used affinity-purification coupled with tandem mass tag-based quantitative mass spectrometry to assess protein interaction changes of WT KV11.1 or trafficking-deficient channel variants in the presence or absence of E-4031. We identified 572 core KV11.1 protein interactors. Trafficking-deficient variants KV11.1-G601S and KV11.1-G601S-G965∗ had significantly increased interactions with proteins responsible for folding, trafficking, and degradation compared to WT. We confirmed previous findings that the proteasome is critical for KV11.1 degradation. Our report provides the first comprehensive characterization of protein quality control mechanisms of KV11.1. We find extensive interactome remodeling associated with trafficking-deficient KV11.1 variants and with pharmacological chaperone rescue of KV11.1 cell surface expression. The identified protein interactions could be targeted therapeutically to improve KV11.1 trafficking and treat LQTS.

The proteostasis interactomes of trafficking-deficient K V 11.1 variants associated with Long QT Syndrome and pharmacological chaperone rescue.

Introduction: The voltage gated potassium ion channel K V 11.1 plays a critical role in cardiac repolarization. Genetic variants that render Kv11.1 dysfunctional cause Long QT Syndrome (LQTS), which is associated with fatal arrhythmias. Approximately 90% of LQTS-associated variants cause intracellular protein transport (trafficking) dysfunction, which can be rescued by pharmacological chaperones like E-4031. Protein folding and trafficking decisions are regulated by chaperones, protein quality control factors, and trafficking machinery, comprising the cellular proteostasis network. Here, we test whether trafficking dysfunction is associated with alterations in the proteostasis network of pathogenic Kv11.1 variants, and whether pharmacological chaperones can normalize the proteostasis network of responsive variants. Methods: We used affinity-purification coupled with tandem mass tag-based quantitative mass spectrometry to assess protein interaction changes in human embryonic kidney (HEK293) cells expressing wild-type (WT) K V 11.1 or trafficking-deficient channel variants in the presence or absence of E-4031. Resultsa: We identified 573 core K V 11.1 protein interactors. Both variants K V 11.1-G601S and K V 11.1-G601S-G965* had significantly increased interactions with proteins responsible for folding, trafficking, and degradation compared to WT. We found that proteasomal degradation is a key component for K V 11.1 degradation and that the K V 11.1-G601S-G965* variant was more responsive to E-4031 treatment. This suggests a role in the C-terminal domain and the ER retention motif of K V 11.1 in regulating trafficking. Conclusion: Our report characterizes the proteostasis network of K V 11.1, two trafficking deficient K V 11.1 variants, and variants treated with a pharmacological chaperone. The identified protein interactions could be targeted therapeutically to improve K V 11.1 trafficking and treat Long QT Syndrome.