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ARS Home » Pacific West Area » Corvallis, Oregon » National Clonal Germplasm Repository » Research » Publications at this Location » Publication #307205
Title: Cryopreservation affects ROS-induced oxidative stress and antioxidant response in Arabidopsis seedlings

Author
item CHEN, GUAN-QUN - Shanghai Jiaotong University
item REN, LI - Shanghai Jiaotong University
item ZHANG, JIE - Shanghai Jiaotong University
item Reed, Barbara
item ZHANG, DI - Shanghai Jiaotong University
item SHEN, XIAO-HUI - Shanghai Jiaotong University

Submitted to: Cryobiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/30/2014
Publication Date: 12/6/2014
Citation: Chen, G., Ren, L., Zhang, J., Reed, B.M., Zhang, D., Shen, X. 2014. Cryopreservation affects ROS-induced oxidative stress and antioxidant response in Arabidopsis seedlings. Cryobiology. 70(1):38-47.

Interpretive Summary: Arabadopsis seedlings were tested to determine the amount of oxidative stress produced during a cryopreservation protocol. Two groups of Arabidopsis thaliana seedlings, one that had no survival after cryopreservation and one that had moderate survival, were tested for indicators of oxidative stress after 48-h or 72-h of germination at five steps of a cryopreservation protocol. Several indicators of stress were studied. Hydrogen peroxide-induced oxidative stress was found at the steps of dehydration and rapid warming in both 48-h seedlings (high survival rate) and 72-h seedlings (no survival). Low survival of 72-h seedlings was associated with severe membrane damage, which was caused by increased oxidative stress and decreased antioxidant activity. Genes for antioxidant activity were active in 48-h seedlings but not in 72-h seedlings. Enhanced antioxidant activity contributed to the high survival rate of 48-h seedlings. This model system indicates that antioxidants levels might be manipulated to improve the recovery of seedlings after cryopreservation.

Technical Abstract: Plant recovery status after cryopreservation by vitrification had a negative relationship to the oxidative stress induced by reactive oxygen species (ROS). Arabidopsis thaliana seedlings germinated for 48-h or 72-h with different cryopreservation survival tolerances were examined at five steps of a vitrification-based cryopreservation protocol, to determine the role of ROS (O2•-, H2O2 and OH•) and antioxidant systems (SOD, POD, CAT, AsA and GSH) in cryo-injury. In addition the effects of the five steps on membrane lipid peroxidation were studied using malondialdehyde (MDA) as an indicator. Correlation analysis indicated that H2O2-induced oxidative stress at the steps of dehydration and rapid warming was the main cause of cryo-injury of 48-h seedlings (high survival rate) and 72-h seedlings (no survival). H2O2 was mainly generated in cotyledons, shoot tips and roots of seedlings as indicated by Amplex Red staining. Low survival of 72-h seedlings was associated with severe membrane lipid peroxidation, which was caused by increased OH• generation activity and decreased SOD activity. The antioxidant-related gene expression by qRT-PCR and physiological assays suggested that the antioxidant systems of 48-h seedlings were activated by ROS, and they actively mounted a defense against oxidative stress during cryopreservation. A high level of ROS led to the weakening of the antioxidant system of 72-h seedlings. Enhanced CAT activities contributed to the high survival rate of 48-h seedlings as indicated by cryopreservation of antioxidant mutant seedlings. This model system indicates that elevated AsA content and CAT activities were determinants of cryogenic stress tolerance, whose manipulation could improve the recovery of seedlings after cryopreservation.