SOD2 targeted gene editing by CRISPR/Cas9 yields Human cells devoid of MnSOD.

Free Radic Biol Med

Department of Radiation Oncology, B180 Medical Laboratories, The University of Iowa, Iowa City, IA 52242. Electronic address:

Published: December 2015

To date no models exist to study MnSOD deficiency in human cells. To address this deficiency, we created a SOD2-null human cell line that is completely devoid of detectable MnSOD protein expression and enzyme activity. We utilized the CRISPR/Cas9 system to generate biallelic SOD2 disruption in HEK293T cells. These SOD2-null cells exhibit impaired clonogenic activity, which was rescued by either treatment with GC4419, a pharmacological small-molecule mimic of SOD, or growth in hypoxia. The phenotype of these cells is primarily characterized by impaired mitochondrial bioenergetics. The SOD2-null cells displayed perturbations in their mitochondrial ultrastructure and preferred glycolysis as opposed to oxidative phosphorylation to generate ATP. The activities of mitochondrial complex I and II were both significantly impaired by the absence of MnSOD activity, presumably from disruption of the Fe/S centers in NADH dehydrogenase and succinate dehydrogenase subunit B by the aberrant redox state in the mitochondrial matrix of SOD2-null cells. By creating this model we provide a novel tool with which to study the consequences of lack of MnSOD activity in human cells.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4890619PMC
http://dx.doi.org/10.1016/j.freeradbiomed.2015.07.017DOI Listing

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