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Cloning and functional analysis of HvERF039 gene in Qingke (Hordeum vulgare L. var. nudum Hook. f)

MA Juan-E, YAO You-Hua, YAO Xiao-Hua, WU Kun-Lun*, and CUI Yong-Mei*   

  1. Academy of Agriculture and Forestry Sciences, Qinghai University / State Key Laboratory of Plateau Ecology and Agriculture, Qinghai University / Laboratory for Research and Utilization of Qinghai Tibet Plateau Germplasm Resources / Qinghai Key Laboratory of Hulless Barley Genetics and Breeding, Xining 810016, Qinghai, China
  • Received:2025-01-24 Revised:2025-06-01 Accepted:2025-06-01 Published:2025-06-19
  • Supported by:
    This study was supported by the Open Project of State Key Laboratory of Plateau Ecology and Agriculture (2023-ZZ-01), the Qinghai University Natural Science Foundation for Young Scholars (2022-QNY-3), the Innovation Fund of Qinghai Academy of Agricultural and Forestry Sciences (2022-NKY-04), and the Laboratory for Research and Utilization of Qinghai Tibet Plateau Germplasm Resources (2025). 

Abstract:

AP2/ERF (APETALA2/ethylene responsive factor) transcription factors play crucial roles in responses to abiotic stresses, but their functions remain poorly understood in Qingke (hulless barley). In this study, we cloned the HvERF039 gene and investigated its role in cold stress response through bioinformatics analysis, qRT-PCR, and heterologous expression in Arabidopsis thaliana. HvERF039 encoded a hydrophilic, unstable protein containing a typical AP2 conserved domain. The promoter region of HvERF039 contained various cis-acting elements, including those responsive to light, hormones and cold stress. Subcellular localization and transactivation assays revealed that HvERF039 protein was localized in both the membrane and nucleus and possessed transcriptional activation activity. qRT-PCR analysis showed that HvERF039 expression was significantly upregulated under cold treatment, with the highest expression observed in leaves and detectable levels in most other tissues. Functional analysis demonstrated that transgenic Arabidopsis plants overexpressing HvERF039 exhibited significantly higher germination and survival rates under cold stress compared to wild-type plants. These transgenic lines also showed enhanced physiological resistance, including lower ion leakage, reduced H2O2 and MDA contents, and increased CAT activity. Furthermore, HvERF039 was found to physically interact with multiple stress-related proteins. Collectively, these findings suggest that HvERF039 plays a positive regulatory role in cold stress tolerance and represents a promising genetic resource for improving cold resistance in Qingke cultivars.

Key words: Qingke, HvERF039, transcription factors, heterologous overexpression, cold stress

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