Developmental Angiogenesis Requires the Mitochondrial Phenylalanyl-tRNA Synthetase

Abstract

Background: Mitochondrial aminoacyl-tRNA synthetases (mtARSs) catalyze the binding of specific amino acids to their cognate tRNAs and play an essential role in the synthesis of proteins encoded by mitochondrial DNA. Defects in mtARSs have been linked to human diseases, but their tissue-specific pathophysiology remains elusive. Here we examined the role of mitochondrial phenylalanyl-tRNA synthetase (FARS2) in developmental angiogenesis and its potential contribution to the pathogenesis of cardiovascular disease.Methods: Morpholinos were injected into fertilized zebrafish ova to establish an in vivo fars2 knock-down model. A visualization of the vasculature was achieved by using Tg (fli1: EGFP)y1 transgenic zebrafish. In addition, small interference RNAs (siRNAs) were transferred into human umbilical vein endothelial cells (HUVECs) to establish an in vitro FARS2 knock-down model. Cell motility, proliferation, and tubulogenesis were determined using scratch-wound CCK8, transwell-based migration, and tube formation assays. In addition, mitochondria- and non-mitochondria-related respiration were evaluated using a Seahorse XF24 analyzer and flow cytometry assays. Analyses of the expression levels of transcripts and proteins were performed using qRT-PCR and western blotting, respectively.Results: The knock-down of fars2 hampered the embryonic development in zebrafish and delayed the formation of the vasculature in Tg (fli1: EGFP)y1 transgenic zebrafish. In addition, the siRNA-mediated knock-down of FARS2 impaired angiogenesis in HUVECs as indicated by decreased cell motility and tube formation capacity. The knock-down of FARS2 also produced variable decreases in mitochondrial- and non-mitochondrial respiration in HUVECs and disrupted the regulatory pathways of angiogenesis in both HUVECs and zebrafish.Conclusion: Our current work offers novel insights into angiogenesis defects and cardiovascular diseases induced by FARS2 deficiency.

Mutations in DARS2 result in global dysregulation of mRNA metabolism and splicing

“…Furthermore, no correlation was found between disease severity and residual enzyme activity of mt-AspRS from LBSL patient lymphoblasts [1]. Housekeeping aminoacylation seems not to be the major target of these mutations, pointing to the possibility that mt-AspRS moonlights in the cells by performing non-canonical functions such as angiogenesis, immune response, tumorigenesis or neurodevelopment, as has been reported for several other aminoacyl-tRNA synthetases [41,42]. Therefore, the possibility that mt-AspRS is directly involved into the regulation of RBPs cannot be ruled out.…”

Mitochondrial UQCC3 controls angiogenesis during embryonic development and tumor growth

“…As others have recently described, many mitochondrial proteins involved in angiogenesis. The knockdown of FARS2 hampered the embryonic development in zebra sh and delayed the formation of the vasculature [32]. In hypoxic pancreatic tumors PKM2 interferes both with NF-κB/p65 and HIF-1α activation that ultimately triggers VEGF-A secretion and subsequent blood vessel formation [33].…”

Disease models of mitochondrial aminoacyl‐tRNA synthetase defects

“…Knockdowns of mtARSs have also been studied in other animal models, particularly in zebrafish and flies. The zebrafish models include FARS2, 25,44 RARS2, 45 VARS2, 46 YARS2, 47 and WARS2 11 knockdowns, with phenotypes ranging from retarded embryonic development to brain hypoplasia, heart failure, and CNS and skeletal muscle involvement. Findings resembling those in mtARS mouse models have been reported in zebrafish models, including respiratory chain deficiency, ISR activation, and disrupted fatty acid oxidation 46 .…”