Altered Activation of Protein Kinase PKR and Enhanced Apoptosis in Dystonia Cells Carrying a Mutation in PKR Activator Protein PACT

• Published in: The Journal of biological chemistry 2015-09, Vol.290 (37), p.22543-22557
• Main Authors: Vaughn, Lauren S, Bragg, D. Cristopher, Sharma, Nutan, Camargos, Sarah, Cardoso, Francisco, Patel, Rekha C
• Format: Article
• Language: English
• Subjects: Amino Acid Substitution; Apoptosis; cell death; Dystonia - genetics; Dystonia - metabolism; Dystonia - pathology; eIF-2 Kinase - genetics; eIF-2 Kinase - metabolism; endoplasmic reticulum stress (ER stress); Female; HeLa Cells; Humans; interferon; Male; Molecular Bases of Disease; Mutation, Missense; protein kinase RNA-activated (PKR); RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M115.669408

PACT is a stress-modulated activator of the interferon-induced double-stranded RNA-activated protein kinase (PKR). Stress-induced phosphorylation of PACT is essential for PACT's association with PKR leading to PKR activation. PKR activation leads to phosphorylation of translation initiation factor eIF2α inhibition of protein synthesis and apoptosis. A recessively inherited form of early-onset dystonia DYT16 has been recently identified to arise due to a homozygous missense mutation P222L in PACT. To examine if the mutant P222L protein alters the stress-response pathway, we examined the ability of mutant P222L to interact with and activate PKR. Our results indicate that the substitution mutant P222L activates PKR more robustly and for longer duration albeit with slower kinetics in response to the endoplasmic reticulum stress. In addition, the affinity of PACT-PACT and PACT-PKR interactions is enhanced in dystonia patient lymphoblasts, thereby leading to intensified PKR activation and enhanced cellular death. P222L mutation also changes the affinity of PACT-TRBP interaction after cellular stress, thereby offering a mechanism for the delayed PKR activation in response to stress. Our results demonstrate the impact of a dystonia-causing substitution mutation on stress-induced cellular apoptosis. Background: Point mutation P222L in PKR activator PACT causes movement disorder dystonia. Results: PACT mutant P222L causes delayed and prolonged PKR and eIF2α phosphorylation in response to the ER stress. Conclusion: Altered kinetics of PKR activation in response to the ER stress enhances apoptosis in dystonia cells. Significance: This is the first study of molecular mechanisms involved in dystonia 16.