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The Heat Stress Transcription Factor LlHsfA4 Enhanced Basic Thermotolerance through Regulating ROS Metabolism in Lilies (Lilium Longiflorum).

ABSTRACT: Heat stress severely affects the annual agricultural production. Heat stress transcription factors (HSFs) represent a critical regulatory juncture in the heat stress response (HSR) of plants. The HsfA1-dependent pathway has been explored well, but the regulatory mechanism of the HsfA1-independent pathway is still under-investigated. In the present research, HsfA4, an important gene of the HsfA1-independent pathway, was isolated from lilies (Lilium longiflorum) using the RACE method, which encodes 435 amino acids. LlHsfA4 contains a typical domain of HSFs and belongs to the HSF A4 family, according to homology comparisons and phylogenetic analysis. LlHsfA4 was mainly expressed in leaves and was induced by heat stress and H₂O₂ using qRT-PCR and GUS staining in transgenic Arabidopsis. LlHsfA4 had transactivation activity and was located in the nucleus and cytoplasm through a yeast one hybrid system and through transient expression in lily protoplasts. Over expressing LlHsfA4 in Arabidopsis enhanced its basic thermotolerance, but acquired thermotolerance was not achieved. Further research found that heat stress could increase H₂O₂ content in lily leaves and reduced H₂O₂ accumulation in transgenic plants, which was consistent with the up-regulation of HSR downstream genes such as Heat stress proteins (HSPs), Galactinol synthase1 (GolS1), WRKY DNA binding protein 30 (WRKY30), Zinc finger of Arabidopsis thaliana 6 (ZAT6) and the ROS-scavenging enzyme Ascorbate peroxidase 2 (APX2). In conclusion, these results indicate that LlHsfA4 plays important roles in heat stress response through regulating the ROS metabolism in lilies.

SUBMITTER: Wang C

PROVIDER: S-EPMC8745440 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The lily, a famous bulbous flower, is seriously affected by high temperatures, which affect their growth and production. To date, the signalling pathways and the molecular mechanisms related to heat response in Lilium have not been elucidated. In this study, a comparative transcriptome analysis was performed in an important thermo-tolerant flower, L. longiflorum, and a thermo-sensitive flower, L. distichum. Lily seedlings were first exposed to heat stress at 42°C for different lengths of time, and the optimal time-points (2 h and 24 h) were selected for RNA sequencing (RNA-seq). Approximately 66.51, 66.21, and 65.36 Mb clean reads were identified from three libraries of L. longiflorum (LL_CK, LL_T2h and LL_T24h, respectively) and 66.18, 66.03, and 65.16 Mb clean reads were obtained from three libraries of L. distichum (LD_CK, LD_T2h and LD_T24h, respectively) after rRNA removing. A total of 34,301 unigenes showed similarity to known proteins in the database NCBI non-redundant protein (NR), Swiss-Prot proteins, InterPro proteins, Clusters of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG). In addition, 1,621 genes were differentially expressed in the overlapping libraries between LL_DEGs and LD_DEGs; of these genes, 352 DEGs were obviously upregulated in L. longiflorum and downregulated in L. distichum during heat stress, including 4-coumarate, CoA ligase (4CL), caffeoyl-CoA O-methyltransferase (CCoAOMT), peroxidase, pathogenesis-related protein 10 family genes (PR10s), 14-3-3 protein, leucine-rich repeat receptor-like protein kinase, and glycine-rich cell wall structural protein-like. These genes were mainly involved in metabolic pathways, phenylpropanoid biosynthesis, plant-pathogen interactions, plant hormone signal transduction, and kinase signalling pathways. Quantitative RT-PCR was performed to validate the expression profiling of these DEGs in RNA-seq data. Taken together, the results obtained in the present study provide a comprehensive sequence resource for the discovery of heat-resistance genes and reveal potential key components that are responsive to heat stress in lilies, which may help to elucidate the heat signal transcription networks and facilitate heat-resistance breeding in lily.

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The Heat Stress Transcription Factor LlHsfA4 Enhanced Basic Thermotolerance through Regulating ROS Metabolism in Lilies (Lilium Longiflorum).

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