As one of the subtropical and tropical fruit trees, Citrus sinensis is sensitive to cold stress. However, most transcription factors (TFs) that regulate cold tolerance in citrus have not yet been reported. A phytochrome-interacting transcription factor (PIF) gene (CsPIF8) in citrus was significantly upregulated under cold stress. Overexpression of CsPIF8 increased cold tolerance in transgenic tomato plants and grapefruit callus, whereas virus-induced gene silencing-mediated suppression of PIF8 increased cold sensitivity in seedlings of Poncirus trifoliata. Superoxide dismutase (SOD) reduces the superoxide anion (O2-) level to enhance cold tolerance in plants. Chromatin immunoprecipitation combined with high-throughput sequencing, yeast one hybrid, electrophoretic mobility shift and dual luciferase assays showed that CsPIF8 directly bound the E-box (CANNTG) of CsSOD promoter and activated the promoter of CsSOD. Furthermore, the expression level of CsSOD and CsSOD activity were significantly increased, whereas the level of O2- was significantly reduced in the transgenic lines. The Poncirus trifoliata seedlings with VIGS-mediated suppression of PIF8 exhibited the opposite effects. These results have shown that CsPIF8 improved cold tolerance in citrus through regulating the expression level of SOD and SOD activity. These findings may provide novel insights into the regulation of PIF8 in the response to cold stress in citrus.

中文翻译：

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光敏色素相互作用转录因子 CsPIF8 通过调节超氧化物歧化酶表达促进柑橘耐寒性

柑橘作为亚热带和热带果树之一，对冷胁迫敏感。然而，大多数调节柑橘耐寒性的转录因子 (TFs) 尚未报道。柑橘中的光敏色素相互作用转录因子 (PIF) 基因 (CsPIF8) 在冷胁迫下显着上调。CsPIF8 的过表达增加了转基因番茄植物和葡萄柚愈伤组织的耐寒性，而病毒诱导的基因沉默介导的 PIF8 抑制增加了枳实幼苗的冷敏感性。超氧化物歧化酶 (SOD) 降低超氧阴离子 (O2-) 水平以增强植物的耐寒性。染色质免疫沉淀结合高通量测序，酵母一杂交，电泳迁移率变化和双荧光素酶测定表明 CsPIF8 直接结合 CsSOD 启动子的 E-box (CANNTG) 并激活 CsSOD 的启动子。此外，CsSOD 的表达水平和 CsSOD 活性显着增加，而 O2- 水平在转基因品系中显着降低。具有 VIGS 介导的 PIF8 抑制的枳实幼苗表现出相反的效果。这些结果表明，CsPIF8 通过调节 SOD 的表达水平和 SOD 活性来提高柑橘的耐寒性。这些发现可能为 PIF8 在柑橘冷胁迫响应中的调节提供新的见解。而O2-水平在转基因品系中显着降低。具有 VIGS 介导的 PIF8 抑制的枳实幼苗表现出相反的效果。这些结果表明，CsPIF8 通过调节 SOD 的表达水平和 SOD 活性来提高柑橘的耐寒性。这些发现可能为 PIF8 在柑橘冷胁迫响应中的调节提供新的见解。而O2-水平在转基因品系中显着降低。具有 VIGS 介导的 PIF8 抑制的枳实幼苗表现出相反的效果。这些结果表明，CsPIF8 通过调节 SOD 的表达水平和 SOD 活性来提高柑橘的耐寒性。这些发现可能为 PIF8 在柑橘冷胁迫响应中的调节提供新的见解。