Transcriptional regulation mechanisms underlying chilling injury (CI) development have been widely investigated in model plants and cold-sensitive fruits, such as banana (Musa acuminata). However, unlike the well-known NAC and WRKY transcription factors (TFs), the function and deciphering mechanism of heat shock factors (HSFs) involving in cold response are still fragmented. Here, we showed that hot water treatment (HWT) alleviated CI in harvested banana fruits accomplishing with reduced reactive oxygen species (ROS) accumulation and increased antioxidant enzyme activities. A cold-inducible but HWT-inhibited HSF, MaHsf24, was identified. Using DNA affinity purification sequencing (DAP-seq) combined with RNA-seq analyses, we found three heat shock protein (HSP) genes (MaHSP23.6, MaHSP70-1.1 and MaHSP70-1.2) and three antioxidant enzyme genes (MaAPX1, MaMDAR4 and MaGSTZ1) were the potential targets of MaHsf24. Subsequent electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation coupled with quantitative PCR (ChIP-qPCR) and dual-luciferase reporter (DLR) analyses demonstrated that MaHsf24 repressed the transcription of these six targets via directly binding to their promoters. Moreover, stably overexpressing MaHsf24 in tomatoes increased cold sensitivity by suppressing the expressions of HSPs and antioxidant enzyme genes, while HWT could recover cold tolerance, maintaining higher levels of HSPs and antioxidant enzyme genes, and activities of antioxidant enzymes. In contrast, transiently silencing MaHsf24 by virus-induced gene silencing (VIGS) in banana peels conferred cold resistance with the upregulation of MaHSPs and antioxidant enzyme genes. Collectively, our findings support the negative role of MaHsf24 in cold tolerance, and unravel a novel regulatory network controlling bananas CI occurrence, concerning MaHsf24-exerted inhibition of MaHSPs and antioxidant enzyme genes.

中文翻译：

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MaHsf24是一种新型负调节剂，通过抑制热休克蛋白和抗氧化酶基因的表达来调节香蕉果实的耐寒性


冷害（CI）发展的转录调控机制已在模型植物和冷敏感水果（例如香蕉（ Musa acuminata ））中得到广泛研究。然而，与众所周知的NAC和WRKY转录因子（TF）不同，涉及冷反应的热休克因子（HSF）的功能和破译机制仍然不完整。在这里，我们表明热水处理（HWT）减轻了收获的香蕉果实中的CI，减少了活性氧（ROS）的积累并增加了抗氧化酶活性。鉴定出一种冷诱导但 HWT 抑制的 HSF，MaHsf24。使用DNA亲和纯化测序（DAP-seq）结合RNA-seq分析，我们发现了三个热休克蛋白（HSP）基因（ MaHSP23.6 、 MaHSP70-1.1和MaHSP70-1.2 ）和三个抗氧化酶基因（ MaAPX1 、 MaMDAR4和MaGSTZ1 ）是MaHsf24的潜在靶标。随后的电泳迁移率变动分析 (EMSA)、染色质免疫沉淀结合定量 PCR (ChIP-qPCR) 和双荧光素酶报告基因 (DLR) 分析表明，MaHsf24 通过直接结合到其启动子来抑制这六个靶标的转录。此外，在番茄中稳定过表达MaHsf24通过抑制HSP和抗氧化酶基因的表达来提高冷敏感性，而HWT可以恢复耐冷性，维持较高水平的HSP和抗氧化酶基因以及抗氧化酶的活性。 相比之下，通过病毒诱导基因沉默（VIGS）在香蕉皮中暂时沉默MaHsf24可通过MaHSP和抗氧化酶基因的上调赋予抗寒性。总的来说，我们的研究结果支持 MaHsf24 在耐冷性中的负面作用，并揭示了控制香蕉 CI 发生的新调控网络，涉及 MaHsf24 对MaHSP和抗氧化酶基因的抑制。