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Functional characterization of isoform A of eukaryotic translation initiation factor 5A (eIF5A)

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Karina Danielle Pereira Thomaz
Total Authors: 1
Document type: Doctoral Thesis
Press: Rio Claro. 2019-10-16.
Institution: Universidade Estadual Paulista (Unesp). Instituto de Biociências. Rio Claro
Defense date:
Advisor: Augusto Ducati Luchessi

The eukaryotic translation initiation factor 5A (eIF5A) is a highly conserved protein in archaeas and eukaryotes. It is characterized by being the only protein known to present the amino acid residue hypusine, produced from a post-translational modification called hypusination, which is essential for its activity. Currently eIF5A is correlated with the beginning, elongation and termination of translation, and its function has been associated with different cellular, physiological and pathological processes. In humans, eIF5A is predicted to be generated from five transcriptional variants (A, B, C, D and X5). Variants B, C, D and X5 encode the 17 kDa canonical protein B, which has been studied in recent decades. However, variant A is expected to encode an alternative 20 kDa isoform (A isoform), which has an additional 30 amino acid sequence in the N-terminal region. In this context, the present study aimed to characterize unpublished molecular events involving the human eIF5A1 isoform A. In this study, we showed that in HeLa cells, isoform A was detected, which was shown to be subject to hypusination. Bioinformatics analysis showed that isoform A has a high probability of targeting mitochondria. Thus, subcellular fractionation techniques have shown that isoform A co-purifies with mitochondria, unlike isoform B. Analysis of gene silencing in HeLa cells using eIF5A1 isoform A-specific siRNA molecules showed that protein depletion is capable of cause apoptosis in cells, as well as decrease viability and cause changes in the cell cycle. In addition, molecular analyzes have shown that this suppression caused mitochondrial dysfunction, which was evidenced by altered expression of genes involved in oxidative metabolism, and increased production of reactive oxygen species (ROS). Gene expression analyzes also showed that isoform A participates in metabolic shift (change from glycolytic to oxidative metabolism and vice versa). Moreover, the increase in respiratory capacity through suppression of isoform A and changes in mitochondrial morphophysiology evidenced by increased mitochondrial fission and number of ridges, suggest a disordered activity of mitochondria through a compensatory mechanism. Collectively, the results of this study indicate that the eIF5A1 isoform A is a key component in mitochondrial function control representing a promising interface for the study of mitochondrial oxidative metabolism and association with the protein synthesis process. (AU)

FAPESP's process: 13/23620-4 - Functional characterization of the translation factor EIF5A isoform A
Grantee:Karina Danielle Pereira
Support Opportunities: Scholarships in Brazil - Doctorate