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SOD2 targeted gene editing by CRISPR/Cas9 yields Human cells devoid of MnSOD
被引:26
作者:

Cramer-Morales, Kimberly
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机构:
Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA

Heer, Collin D.
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Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA

Mapuskar, Kranti A.
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h-index: 0
机构:
Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA

Domann, Frederick E.
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h-index: 0
机构:
Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA
机构:
[1] Univ Iowa, Dept Radiat Oncol, Med Labs B180, Iowa City, IA 52242 USA
关键词:
Superoxide dismutase;
Mitochondria;
Respiration;
Electron transport;
Iron metabolism;
MANGANESE-SUPEROXIDE-DISMUTASE;
2 DISTINCT REGIONS;
OXIDATIVE STRESS;
BREAST-CANCER;
MUTANT MICE;
ESCHERICHIA-COLI;
MESSENGER-RNA;
LONG ARM;
EXPRESSION;
DELETION;
D O I:
10.1016/j.freeradbiomed.2015.07.017
中图分类号:
Q5 [生物化学];
Q7 [分子生物学];
学科分类号:
071010 ;
081704 ;
摘要:
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. (C) 2015 Elsevier Inc. All rights reserved.
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页码:379 / 386
页数:8
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