Reverse phase protein array reveals correlation of retinoic acid metabolism with cardiomyopathy in Friedreich's Ataxia

Academic Article

Abstract

  • Identifying biomarkers is important for assessment of disease progression, prediction of symptom development, and determination of treatment effectiveness. While unbiased analyses of differential gene expression using next-generation sequencing methods are now routinely conducted, proteomics studies are more challenging because of traditional methods predominantly being low throughput and offering a limited dynamic range for simultaneous detection of hundreds of proteins that drastically differ in their intracellular abundance. We utilized a sensitive and high-throughput proteomic technique, reverse phase protein array (RPPA), to attain protein expression profiles of primary fibroblasts obtained from patients with Friedreich's ataxia (FRDA) and unaffected controls (CTRLs). The RPPA was designed to detect 217 proteins or phosphorylated proteins by individual antibody, and the specificity of each antibody was validated prior to the experiment. Among 62 fibroblast samples (44 FRDA and 18 CTRLs) analyzed, 30 proteins/ phosphoproteins were significantly changed in FRDA fibroblasts compared with CTRL cells (p < 0.05), mostly representing signaling molecules and metabolic enzymes. As expected, frataxin was significantly downregulated in FRDA samples, thus serving as an internal CTRL for assay integrity. Extensive bioinformatics analyses were conducted to correlate differentially expressed proteins with critical disease parameters (e.g., selected symptoms, age of onset, guanine–adenine–adenine sizes, frataxin levels, and Functional Assessment Rating Scale scores). Members of the integrin family of proteins specifically associated with hearing loss in FRDA. Also, RPPA data, combined with results of transcriptome profiling, uncovered defects in the retinoic acid metabolism pathway in FRDA samples. Moreover, expression of aldehyde dehydrogenase family 1 member A3 differed significantly between cardiomyopathy-positive and cardiomyopathy-negative FRDA cohorts, demonstrating that metabolites such as retinol, retinal, or retinoic acid could become potential predictive biomarkers of cardiac presentation in FRDA.
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    Digital Object Identifier (doi)

    Author List

  • Napierala JS; Rajapakshe K; Clark A; Chen YY; Huang S; Mesaros C; Xu P; Blair IA; Hauser LA; Farmer J
  • Volume

  • 20