Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses

Cold stress is a significant factor limiting plant growth and development. Pomegranate is particularly susceptible to low temperatures. Calmodulin-binding transcriptional activators (CAMTAs) are key regulators of cold stress tolerance in plants. In this study, we conducted a comprehensive analysis o...

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Main Authors: Shuangshuang Zhao, Rui Lu, Lijuan Feng, Mengyu Zheng, Han Zhang, Yanlei Yin, Ling Zheng
Format: Article
Language:English
Published: MDPI AG 2025-03-01
Series:Plants
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Online Access:https://www.mdpi.com/2223-7747/14/5/813
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author Shuangshuang Zhao
Rui Lu
Lijuan Feng
Mengyu Zheng
Han Zhang
Yanlei Yin
Ling Zheng
author_facet Shuangshuang Zhao
Rui Lu
Lijuan Feng
Mengyu Zheng
Han Zhang
Yanlei Yin
Ling Zheng
author_sort Shuangshuang Zhao
collection DOAJ
description Cold stress is a significant factor limiting plant growth and development. Pomegranate is particularly susceptible to low temperatures. Calmodulin-binding transcriptional activators (CAMTAs) are key regulators of cold stress tolerance in plants. In this study, we conducted a comprehensive analysis of the CAMTA family proteins across 12 species, including <i>Punica granatum</i> (pomegranate), using bioinformatic methods. Pomegranate <i>CAMTA3</i> (<i>PgCAMTA3</i>) was isolated and characterized, and it demonstrated enhanced cold tolerance when expressed in <i>Arabidopsis thaliana</i>. Quantitative real-time PCR (qRT-PCR) analysis showed that the expression of <i>PgCAMTA3</i> was up-regulated under cold and ABA treatments in pomegranates. Two <i>A. thaliana</i> transgenic lines, OE1 and OE2, which overexpress PgCAMTA3, were generated through genetic transformation. The overexpression of <i>PgCAMTA3</i> enhanced the cold stress tolerance in transgenic <i>A. thaliana</i>. OE1 and OE2 exhibited higher survival rates under cold stress. Furthermore, enzymatic activity assays revealed enhanced peroxidase (POD), catalase (CAT), and superoxide dismutase (SOD) in OE lines. These antioxidant enzymatic activities collectively contribute to better cold stress tolerance by providing more effective reactive oxygen species (ROS) scavenging and cellular protection mechanisms, which was confirmed by lower levels of malondialdehyde (MDA) and ROS production. In addition, the overexpression of <i>PgCAMTA3</i> led to the upregulation of the expression levels of <i>AtCBF2</i>, <i>AtNCED3</i>, and <i>AtWRKY22</i>, which were modulated by <i>CAMTA3</i>. In summary, we report the significant role of <i>PgCAMTA3</i> in plant cold tolerance. Our findings provide valuable insights into the CAMATA family in plants and offer new perspectives on the molecular mechanisms underlying cold tolerance in pomegranates.
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spelling doaj-art-eaca0cfeec3e4cb5bc5101eb6a58559b2025-08-20T02:53:19ZengMDPI AGPlants2223-77472025-03-0114581310.3390/plants14050813Functional Characterization of Pomegranate CAMTA3 in Cold Stress ResponsesShuangshuang Zhao0Rui Lu1Lijuan Feng2Mengyu Zheng3Han Zhang4Yanlei Yin5Ling Zheng6Shandong Provincial Key Laboratory of Plant Stress, Life Science College, Shandong Normal University, Jinan 250014, ChinaShandong Provincial Key Laboratory of Plant Stress, Life Science College, Shandong Normal University, Jinan 250014, ChinaShandong Institute of Pomology, Taian 271000, ChinaShandong Provincial Key Laboratory of Plant Stress, Life Science College, Shandong Normal University, Jinan 250014, ChinaShandong Provincial Key Laboratory of Plant Stress, Life Science College, Shandong Normal University, Jinan 250014, ChinaShandong Institute of Pomology, Taian 271000, ChinaShandong Provincial Key Laboratory of Plant Stress, Life Science College, Shandong Normal University, Jinan 250014, ChinaCold stress is a significant factor limiting plant growth and development. Pomegranate is particularly susceptible to low temperatures. Calmodulin-binding transcriptional activators (CAMTAs) are key regulators of cold stress tolerance in plants. In this study, we conducted a comprehensive analysis of the CAMTA family proteins across 12 species, including <i>Punica granatum</i> (pomegranate), using bioinformatic methods. Pomegranate <i>CAMTA3</i> (<i>PgCAMTA3</i>) was isolated and characterized, and it demonstrated enhanced cold tolerance when expressed in <i>Arabidopsis thaliana</i>. Quantitative real-time PCR (qRT-PCR) analysis showed that the expression of <i>PgCAMTA3</i> was up-regulated under cold and ABA treatments in pomegranates. Two <i>A. thaliana</i> transgenic lines, OE1 and OE2, which overexpress PgCAMTA3, were generated through genetic transformation. The overexpression of <i>PgCAMTA3</i> enhanced the cold stress tolerance in transgenic <i>A. thaliana</i>. OE1 and OE2 exhibited higher survival rates under cold stress. Furthermore, enzymatic activity assays revealed enhanced peroxidase (POD), catalase (CAT), and superoxide dismutase (SOD) in OE lines. These antioxidant enzymatic activities collectively contribute to better cold stress tolerance by providing more effective reactive oxygen species (ROS) scavenging and cellular protection mechanisms, which was confirmed by lower levels of malondialdehyde (MDA) and ROS production. In addition, the overexpression of <i>PgCAMTA3</i> led to the upregulation of the expression levels of <i>AtCBF2</i>, <i>AtNCED3</i>, and <i>AtWRKY22</i>, which were modulated by <i>CAMTA3</i>. In summary, we report the significant role of <i>PgCAMTA3</i> in plant cold tolerance. Our findings provide valuable insights into the CAMATA family in plants and offer new perspectives on the molecular mechanisms underlying cold tolerance in pomegranates.https://www.mdpi.com/2223-7747/14/5/813pomegranatecold stress<i>CAMTA</i> genes
spellingShingle Shuangshuang Zhao
Rui Lu
Lijuan Feng
Mengyu Zheng
Han Zhang
Yanlei Yin
Ling Zheng
Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses
Plants
pomegranate
cold stress
<i>CAMTA</i> genes
title Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses
title_full Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses
title_fullStr Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses
title_full_unstemmed Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses
title_short Functional Characterization of Pomegranate CAMTA3 in Cold Stress Responses
title_sort functional characterization of pomegranate camta3 in cold stress responses
topic pomegranate
cold stress
<i>CAMTA</i> genes
url https://www.mdpi.com/2223-7747/14/5/813
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