Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis

In addition to its canonical role in aminoacylation, threonyl-tRNA synthetase (TARS) possesses pro-angiogenic activity that is susceptible to the TARS-specific antibiotic borrelidin. However, the therapeutic benefit of borrelidin is offset by its strong toxicity to living cells. The removal of a sin...

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Main Author: Mirando, Adam Christopher
Format: Others
Language:en
Published: ScholarWorks @ UVM 2015
Subjects:
Online Access:http://scholarworks.uvm.edu/graddis/523
http://scholarworks.uvm.edu/cgi/viewcontent.cgi?article=1522&context=graddis
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spelling ndltd-uvm.edu-oai-scholarworks.uvm.edu-graddis-15222017-03-17T08:44:52Z Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis Mirando, Adam Christopher In addition to its canonical role in aminoacylation, threonyl-tRNA synthetase (TARS) possesses pro-angiogenic activity that is susceptible to the TARS-specific antibiotic borrelidin. However, the therapeutic benefit of borrelidin is offset by its strong toxicity to living cells. The removal of a single methylene group from the parent borrelidin generates BC194, a modified compound with significantly reduced toxicity but comparable anti-angiogenic potential. Biochemical analyses revealed that the difference in toxicities was due to borrelidin's stimulation of amino acid starvation at ten-fold lower concentrations than BC194. However, both compounds were found to inhibit in vitro and in vivo models of angiogenesis at sub-toxic concentrations, suggesting a similar mechanism that is distinct from the toxic responses. Crystal structures of TARS in complex with each compound indicated that the decreased contacts in the BC194 structure may render it more susceptible to competition with the canonical substrates and permit sufficient aminoacylation activity over a wider concentration of inhibitor. Conversely, both borrelidin and BC194 induce identical conformational changes in TARS, providing a rationale for their comparable effects on angiogenesis. The mechanisms of TARS and borrelidin-based compounds on angiogenesis were subsequently tested using zebrafish and cell-based models. These data revealed ectopic branching, non-functional vessels, and increased cell-cell contracts following BC194-treatment or knockdown of TARS expression, suggesting a role for the enzyme in the maturation and guidance of nascent vasculature. Using various TARS constructs this function was found to be dependent on two interactions or activities associated with the TARS enzyme that are distinct from its canonical aminoacylation activity. Furthermore, observations that TARS may influence VEGF expression and purinergic signaling suggest the possibility for a receptor-mediated response. Taken together, the results presented here demonstrate a clear role for TARS in angiogenesis, independent of its primary function in translation. Although the exact molecular mechanisms through which TARS and borrelidin regulate this activity remain to be determined, these data provide a foundation for future investigations of TARS's function in vascular biology and its use as a target for angiogenesis-based therapy. 2015-01-01T08:00:00Z text application/pdf http://scholarworks.uvm.edu/graddis/523 http://scholarworks.uvm.edu/cgi/viewcontent.cgi?article=1522&context=graddis Graduate College Dissertations and Theses en ScholarWorks @ UVM Aminoacyl-tRNA Synthetase Angiogenesis Endothelial Cells Enzyme Inhibition Non-canonical Function Polyketide Biochemistry Cell Biology Pharmacology
collection NDLTD
language en
format Others
sources NDLTD
topic Aminoacyl-tRNA Synthetase
Angiogenesis
Endothelial Cells
Enzyme Inhibition
Non-canonical Function
Polyketide
Biochemistry
Cell Biology
Pharmacology
spellingShingle Aminoacyl-tRNA Synthetase
Angiogenesis
Endothelial Cells
Enzyme Inhibition
Non-canonical Function
Polyketide
Biochemistry
Cell Biology
Pharmacology
Mirando, Adam Christopher
Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis
description In addition to its canonical role in aminoacylation, threonyl-tRNA synthetase (TARS) possesses pro-angiogenic activity that is susceptible to the TARS-specific antibiotic borrelidin. However, the therapeutic benefit of borrelidin is offset by its strong toxicity to living cells. The removal of a single methylene group from the parent borrelidin generates BC194, a modified compound with significantly reduced toxicity but comparable anti-angiogenic potential. Biochemical analyses revealed that the difference in toxicities was due to borrelidin's stimulation of amino acid starvation at ten-fold lower concentrations than BC194. However, both compounds were found to inhibit in vitro and in vivo models of angiogenesis at sub-toxic concentrations, suggesting a similar mechanism that is distinct from the toxic responses. Crystal structures of TARS in complex with each compound indicated that the decreased contacts in the BC194 structure may render it more susceptible to competition with the canonical substrates and permit sufficient aminoacylation activity over a wider concentration of inhibitor. Conversely, both borrelidin and BC194 induce identical conformational changes in TARS, providing a rationale for their comparable effects on angiogenesis. The mechanisms of TARS and borrelidin-based compounds on angiogenesis were subsequently tested using zebrafish and cell-based models. These data revealed ectopic branching, non-functional vessels, and increased cell-cell contracts following BC194-treatment or knockdown of TARS expression, suggesting a role for the enzyme in the maturation and guidance of nascent vasculature. Using various TARS constructs this function was found to be dependent on two interactions or activities associated with the TARS enzyme that are distinct from its canonical aminoacylation activity. Furthermore, observations that TARS may influence VEGF expression and purinergic signaling suggest the possibility for a receptor-mediated response. Taken together, the results presented here demonstrate a clear role for TARS in angiogenesis, independent of its primary function in translation. Although the exact molecular mechanisms through which TARS and borrelidin regulate this activity remain to be determined, these data provide a foundation for future investigations of TARS's function in vascular biology and its use as a target for angiogenesis-based therapy.
author Mirando, Adam Christopher
author_facet Mirando, Adam Christopher
author_sort Mirando, Adam Christopher
title Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis
title_short Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis
title_full Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis
title_fullStr Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis
title_full_unstemmed Characterization Of A Non-Canonical Function For Threonyl-Trna Synthetase In Angiogenesis
title_sort characterization of a non-canonical function for threonyl-trna synthetase in angiogenesis
publisher ScholarWorks @ UVM
publishDate 2015
url http://scholarworks.uvm.edu/graddis/523
http://scholarworks.uvm.edu/cgi/viewcontent.cgi?article=1522&context=graddis
work_keys_str_mv AT mirandoadamchristopher characterizationofanoncanonicalfunctionforthreonyltrnasynthetaseinangiogenesis
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