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Biochem J. 2019 Jun 11;476(11):1637-1651. doi: 10.1042/BCJ20190040.

Characterization of soluble CD39 (SolCD39/NTPDase1) from PiggyBac nonviral system as a tool to control the nucleotides level.

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Laboratory of Cell Biology, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, Brazil.
Department of Biophysics and Center of Biotechnology, Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, RS, Brazil.
Département de Microbiologie-Infectiologie et d'Immunologie, Faculté de Médecine, Université Laval, Québec City, QC G1V 0A6, Canada.
Centre de recherche du CHU de Québec-Université Laval, Québec City, QC G1V 4G2, Canada.
Laboratory of Cell Biology, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, Brazil


Extracellular ATP (eATP) and its metabolites have emerged as key modulators of different diseases and comprise a complex pathway called purinergic signaling. An increased number of tools have been developed to study the role of nucleotides and nucleosides in cell proliferation and migration, influence on the immune system and tumor progression. These tools include receptor agonists/antagonists, engineered ectonucleotidases, interference RNAs and ectonucleotidase inhibitors that allow the control and quantification of nucleotide levels. NTPDase1 (also called apyrase, ecto-ATPase and CD39) is one of the main enzymes responsible for the hydrolysis of eATP, and purified enzymes, such as apyrase purified from potato, or engineered as soluble CD39 (SolCD39), have been widely used in in vitro and in vivo experiments. However, the commercial apyrase had its effects recently questioned and SolCD39 exhibits limitations, such as short half-life and need of high doses to reach the expected enzymatic activity. Therefore, this study investigated a non-viral method to improve the overexpression of SolCD39 and evaluated its impact on other enzymes of the purinergic system. Our data demonstrated that PiggyBac transposon system proved to be a fast and efficient method to generate cells stably expressing SolCD39, producing high amounts of the enzyme from a limited number of cells and with high hydrolytic activity. In addition, the soluble form of NTPDase1/CD39 did not alter the expression or catalytic activity of other enzymes from the purinergic system. Altogether, these findings set the groundwork for prospective studies on the function and therapeutic role of eATP and its metabolites in physiological and pathological conditions.


CD39; PiggyBac; apyrase; purinergic signaling; soluble NTPDase1; transposon


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