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<p>The C2 domain-containing protein family is pivotal for calcium-dependent signaling, membrane trafficking, and abiotic stress adaptation in plants. Despite its importance, a comprehensive genome-wide characterization of this family in <i>Populus deltoides,</i> a species of significant economic and ecological, has not yet been conducted. Here, a total of 115 C2 members were identified in <i>P. deltoides</i> genome, characterized by conserved physicochemical properties, distinct phylogenetic clades, uneven chromosomal distribution, and extensive segmental duplication. Promoter sequence analysis uncovered numerous cis-regulatory elements associated with hormone signaling, stress responsiveness, light-responsive, and growth and development processes. Expression profiling under PEG6000-simulated drought stress revealed divergent expression patterns across the PdeC2 family. Through weighted gene co-expression network analysis (WGCNA), we delineated a specific drought-responsive module containing 38 PdeC2 genes, identifying <i>PdeC2-65</i> as a core hub gene, alongside with <i>PdeC2-81</i> and <i>PdeC2-22</i>. Furthermore, subnetwork analysis pinpointed 16 PdeC2 genes significantly upregulated by PEG, mannitol, and abscisic acid (ABA). Subcellular localization analysis revealed that PdeC2-65 resides primarily in the plasma membrane, whereas PdeC2-38 and PdeC2-57 localize to both the plasma membrane and nucleus. Crucially, yeast two-hybrid (Y2H) and split-luciferase (Split-LUC) assays validated physical interactions among PdeC2-65, PdeC2-38, and PdeC2-57. These interactions were markedly enhanced by Ca<sup>2</sup>⁺ treatment, indicating that these proteins likely assemble into a calcium-dependent functional complex to regulate stress responses. Transient transformation assays further confirmed that overexpression of <i>PdeC2-57</i> or <i>PdeC2-65</i> significantly enhances osmotic stress tolerance in <i>P. deltoides</i>, alleviates membrane lipid peroxidation damage, and activates the reactive oxygen species scavenging system. Collectively, our study provides the first systematic genomic analysis of C2 family in <i>P. deltoids</i>, offering new insights into the molecular mechanisms of PdeC2-mediated osmotic stress tolerance and providing valuable genetic resources for breeding drought-resilient poplar varieties.</p>

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Genome-Wide Identification and Functional Characterization of the C2 Gene Family in Populus deltoides: A PdeC2-38/57/65 Complex Mediates Osmotic Stress Response via Ca2+ Signaling

  • 期刊论文

  • 发表日期：

    2026年5月21日

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  • Xun Cao,
  • Zhengquan He,
  • Hu Huang,
  • Tianheng Jiang,
  • Juan Chen,
  • Anqi Chen,
  • Daobin Ji,
  • Yu Chen,
  • Feifei Lu,
  • Renying Zhuo,
  • Jing Xu

摘要

The C2 domain-containing protein family is pivotal for calcium-dependent signaling, membrane trafficking, and abiotic stress adaptation in plants. Despite its importance, a comprehensive genome-wide characterization of this family in Populus deltoides, a species of significant economic and ecological, has not yet been conducted. Here, a total of 115 C2 members were identified in P. deltoides genome, characterized by conserved physicochemical properties, distinct phylogenetic clades, uneven chromosomal distribution, and extensive segmental duplication. Promoter sequence analysis uncovered numerous cis-regulatory elements associated with hormone signaling, stress responsiveness, light-responsive, and growth and development processes. Expression profiling under PEG6000-simulated drought stress revealed divergent expression patterns across the PdeC2 family. Through weighted gene co-expression network analysis (WGCNA), we delineated a specific drought-responsive module containing 38 PdeC2 genes, identifying PdeC2-65 as a core hub gene, alongside with PdeC2-81 and PdeC2-22. Furthermore, subnetwork analysis pinpointed 16 PdeC2 genes significantly upregulated by PEG, mannitol, and abscisic acid (ABA). Subcellular localization analysis revealed that PdeC2-65 resides primarily in the plasma membrane, whereas PdeC2-38 and PdeC2-57 localize to both the plasma membrane and nucleus. Crucially, yeast two-hybrid (Y2H) and split-luciferase (Split-LUC) assays validated physical interactions among PdeC2-65, PdeC2-38, and PdeC2-57. These interactions were markedly enhanced by Ca2⁺ treatment, indicating that these proteins likely assemble into a calcium-dependent functional complex to regulate stress responses. Transient transformation assays further confirmed that overexpression of PdeC2-57 or PdeC2-65 significantly enhances osmotic stress tolerance in P. deltoides, alleviates membrane lipid peroxidation damage, and activates the reactive oxygen species scavenging system. Collectively, our study provides the first systematic genomic analysis of C2 family in P. deltoids, offering new insights into the molecular mechanisms of PdeC2-mediated osmotic stress tolerance and providing valuable genetic resources for breeding drought-resilient poplar varieties.