Stress and damage mechanisms in Dendrobium nobile Lindl. protocorm-like bodies during pre- and post-liquid nitrogen exposure in cryopreservation revealed by iTRAQ proteomic analysis

Abstract

Liquid nitrogen (LN) exposure, a crucial step in cryopreservation, is mainly responsible for the reduced viability of protocorm-like bodies (PLBs) in Dendrobium nobile Lindl. To characterize the stress and damage mechanisms during pre- and post-LN exposure, the isobaric tags for relative and absolute quantification (iTRAQ)-labeling method was used to evaluate proteome profiles of PLBs in D. nobile ‘Hamana Lake Dream’ undergoing the cooling and rewarming process. In total, 1747 proteins were detected; a cluster of orthologous group (COG) analysis of the identified proteins revealed that posttranslational modification, protein turnover, and chaperone-related proteins were the predominant protein classes. Of these, 108 proteins were differentially changed and grouped into different functional categories, which included protein turnover, stress and defense, carbohydrate and energy metabolism, signaling transduction, metabolism, membrane, and transport. Furthermore, a quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that some proteins, such as annexin, l-ascorbate peroxidase 1, copper/zinc superoxide dismutase, glutathione S-transferase dehydroascorbate reductase 2-like, and an adenosine triphosphate (ATP) synthase, were regulated at their transcriptional levels. Based on functional analysis of these proteins, it was found that protein synthesis, processing, and degradation might be the main strategies used to reestablish cell balance after the cooling and rewarming process. Moreover, the production of reactive oxygen species (ROS) and the decline in energy production, signaling transduction, and membrane transport during pre- and post-LN exposure might be responsible for the viability loss of the PLBs. This work provides potential protein candidates for exploring the stress and cryo-injury mechanism in cryopreservation.