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Ectopic expression of a novel cold-resistance protein 1 from Brassica oleracea promotes tolerance to chilling stress in transgenic tomato.

ABSTRACT: Cold stress is considered as one of the major environmental factors that adversely affects the plant growth and distribution. Therefore, there arises an immediate need to cultivate effective strategies aimed at developing stress-tolerant crops that would boost the production and minimise the risks associated with cold stress. In this study, a novel cold-responsive protein1 (BoCRP1) isolated from Brassica oleracea was ectopically expressed in a cold susceptible tomato genotype Shalimar 1 and its function was investigated in response to chilling stress. BoCRP1 was constitutively expressed in all the tissues of B. oleracea including leaf, root and stem. However, its expression was found to be significantly increased in response to cold stress. Moreover, transgenic tomato plants expressing BoCRP1 exhibited increased tolerance to chilling stress (4 °C) with an overall improved rate of seed germination, increased root length, reduced membrane damage and increased accumulation of osmoprotectants. Furthermore, we observed increased transcript levels of stress responsive genes and enhanced accumulation of reactive oxygen species scavenging enzymes in transgenic plants on exposure to chilling stress. Taken together, these results strongly suggest that BoCRP1 is a promising candidate gene to improve the cold stress tolerance in tomato.

SUBMITTER: Wani UM

PROVIDER: S-EPMC8367951 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundRapid-cycling Brassica napus (B. napus-RC) has potential as a rapid trait testing system for canola (B. napus) because its life cycle is completed within 2 months while canola usually takes 4 months, and it is susceptible to the same range of diseases and abiotic stress as canola. However, a rapid trait testing system for canola requires the development of an efficient transformation and tissue culture system for B. napus-RC. Furthermore, effectiveness of this system needs to be demonstrated by showing that a particular trait can be rapidly introduced into B. napus-RC plants.ResultsAn in-vitro regeneration protocol was developed for B. napus-RC using 4-day-old cotyledons as the explant. High regeneration percentages, exceeding 70%, were achieved when 1-naphthaleneacetic acid (0.10 mg/L), 6-benzylaminopurine (1.0 mg/L), gibberellic acid (0.01 mg/L) and the ethylene antagonist silver nitrate (5 mg/L) were included in the regeneration medium. An average transformation efficiency of 16.4% was obtained using Agrobacterium-mediated transformation of B. napus-RC cotyledons using Agrobacterium strain GV3101 harbouring a plasmid with an NPTII (kanamycin-selectable) marker gene and the Arabidopsis thaliana cDNA encoding ACYL-COA-BINDING PROTEIN6 (AtACBP6). Transgenic B. napus-RC overexpressing AtACBP6 displayed better tolerance to freezing/frost than the wild type, with enhanced recovery from cellular membrane damage at both vegetative and flowering stages. AtACBP6-overexpressing B. napus-RC plants also exhibited lower electrolyte leakage and improved recovery following frost treatment, resulting in higher yields than the wild type. Ovules from transgenic AtACBP6 lines were better protected from frost than those of the wild type, while the developing embryos of frost-treated AtACBP6-overexpressing plants showed less freezing injury than the wild type.ConclusionsThis study demonstrates that B. napus-RC can be successfully regenerated and transformed from cotyledon explants and has the potential to be an effective trait testing platform for canola. Additionally, AtACBP6 shows potential for enhancing cold tolerance in canola however, larger scale studies will be required to further confirm this outcome.

Project description:Storage at low temperatures is a common practice to prolong postharvest life of fruit and vegetables with a minimal negative impact on human/environmental health. Storage at low temperatures, however, can be restricted due to produce susceptibility to non-freezing chilling temperatures, when injuries such as physiological disorders and decays may result in unmarketable produce. We have investigated tomato fruit response to postharvest chilling stress in a recombinant inbred line (RIL) population developed from a cross between a chilling-sensitive cultivated tomato (Solanum lycopersicum L.) breeding line and a chilling-tolerant inbred accession of the tomato wild species S. pimpinellifolium L. Screening of the fruit of 148 RILs under cold storage (1.5°C) indicated presence of significant variations in chilling tolerance, manifested by varying degrees of fruit injury. Two extremely contrasting groups of RILs were identified, chilling-tolerant and chilling-sensitive RILs. The RILs in the two groups were further investigated under chilling stress conditions, and several physiological parameters, including weight loss, chlorophyll fluorescence parameters Fv/Fm, and Performance Index (PI), were determined to be efficient markers for identifying response to chilling stress in postharvest fruit. The Fv/Fm values reflected the physiological damages endured by the fruit after cold storage, and PI was a sensitive marker for early changes in photosystem II function. These two parameters were early indicators of chilling response before occurrence of visible chilling injuries. Antioxidant activities and ascorbic acid content were significantly higher in the chilling-tolerant than the chilling-sensitive lines. Further, the expression of C-repeat/DREB binding factors (CBFs) genes swiftly changed within 1-hr of fruit exposure to the chilling temperature, and the SlCBF1 transcript level was generally higher in the chilling-tolerant than chilling-sensitive lines after 2-hr exposure to the low temperature. This research demonstrates the presence of potential genetic variation in fruit chilling tolerance in the tomato RIL population. Further investigation of the RIL population is underway to better understand the genetic, physiological, and biochemical mechanisms involved in postharvest fruit chilling tolerance in tomato.

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Ectopic expression of a novel cold-resistance protein 1 from Brassica oleracea promotes tolerance to chilling stress in transgenic tomato.

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