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HEAT SHOCK TRANSCRIPTION FACTOR B2b acts as a transcriptional repressor of VIN3, a gene induced by long-term cold for flowering.

ABSTRACT: Vernalization, an acceleration of flowering after long-term winter cold, is an intensively studied flowering mechanism in winter annual plants. In Arabidopsis, Polycomb Repressive Complex 2 (PRC2)-mediated suppression of the strong floral repressor, FLOWERING LOCUS C (FLC), is critical for vernalization and a PHD finger domain protein, VERNALIZATION INSENSITIVE 3 (VIN3), recruits PRC2 on FLC chromatin. The level of VIN3 was found to gradually increase in proportion to the length of cold period during vernalization. However, how plants finely regulate VIN3 expression according to the cold environment has not been completely elucidated. As a result, we performed EMS mutagenesis using a transgenic line with a minimal promoter of VIN3 fused to the GUS reporter gene, and isolated a mutant, hyperactivation of VIN3 1 (hov1), which showed increased GUS signal and endogenous VIN3 transcript levels. Using positional cloning combined with whole-genome resequencing, we found that hov1 carries a nonsense mutation, leading to a premature stop codon on the HEAT SHOCK TRANSCRIPTION FACTOR B2b (HsfB2b), which encodes a repressive heat shock transcription factor. HsfB2b directly binds to the VIN3 promoter, and HsfB2b overexpression leads to reduced acceleration of flowering after vernalization. Collectively, our findings reveal a novel fine-tuning mechanism to regulate VIN3 for proper vernalization response.

SUBMITTER: Jeong G

PROVIDER: S-EPMC9243095 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe effects of diverse stresses ultimately alter the structures and functions of proteins. As molecular chaperones, heat shock proteins (HSPs) are a group of highly conserved proteins that help in the refolding of misfolded proteins and the elimination of irreversibly damaged proteins. They are mediated by a family of transcription factors called heat shock factors (HSFs). The small abalone Haliotis diversicolor is a species naturally distributed along the southern coast of China. In this study, the expression of HdHSF1 was inhibited by RNAi in hemocytes in order to further elucidate the regulatory roles of HdHSF1 on heat shock responsive genes in abalone. Meanwhile, to understand the transcriptional regulation of the HdHSF1 gene, the 5'-upstream regulatory region of HdHSF1 was characterized, and the relative promoter activity was examined by dual-luciferase reporter gene assay system in HEK293T cell lines.ResultsAfter the inhibition of the H. diversicolor HSF1 gene (HdHSF1) by dsRNA (double-stranded RNA), the expression of most heat shock related-genes was down-regulated (p < 0.05). It indicated the importance of HdHSF1 in the heat shock response of H. diversicolor. Meanwhile, 5'-flanking region sequence (2633 bp) of the HdHSF1 gene was cloned; it contained a putative core promoter region, TATA box, CAAT box, CpG island, and many transcription elements. In HEK293T cells, the 5'-flanking region sequence can drive expression of the enhanced green fluorescent protein (EGFP), proving its promoter function. Exposure of cells to the high-temperature (39 °C and 42 °C) resulted in the activation of HdHSF1 promoter activity, which may explain why the expression of the HdHSF1 gene participates in heat shock response. Luciferase activity of different recombinant plasmids, which contained different truncated promoter fragments of the HdHSF1 gene in HEK293T cells, revealed the possible active regions of the promoter. To further identify the binding site of the critical transcription factor in the region, an expression vector with the site-directed mutation was constructed. After being mutated on the GATA-1 binding site, we found that the luciferase activity was significantly increased, which suggested that the GATA-1 binding site has a certain weakening effect on the activity of the HdHSF1 promoter.ConclusionsThese findings suggest that GATA-1 may be one of the transcription factors of HdHSF1, and a possible signaling pathway mediated by HdHSF1 may exist in H. diversicolor to counteract the adverse effects of heat shock stress.

Project description:BackgroundThe heat shock response of Arabidopsis thaliana is dependent upon a complex regulatory network involving twenty-one known transcription factors and four heat shock protein families. It is known that heat shock proteins (Hsps) and transcription factors (Hsfs) are involved in cellular response to various forms of stress besides heat. However, the role of Hsps and Hsfs under cold and non-thermal stress conditions is not well understood, and it is unclear which types of stress interact least and most strongly with Hsp and Hsf response pathways. To address this issue, we have analyzed transcriptional response profiles of Arabidopsis Hsfs and Hsps to a range of abiotic and biotic stress treatments (heat, cold, osmotic stress, salt, drought, genotoxic stress, ultraviolet light, oxidative stress, wounding, and pathogen infection) in both above and below-ground plant tissues.ResultsAll stress treatments interact with Hsf and Hsp response pathways to varying extents, suggesting considerable cross-talk between heat and non-heat stress regulatory networks. In general, Hsf and Hsp expression was strongly induced by heat, cold, salt, and osmotic stress, while other types of stress exhibited family or tissue-specific response patterns. With respect to the Hsp20 protein family, for instance, large expression responses occurred under all types of stress, with striking similarity among expression response profiles. Several genes belonging to the Hsp20, Hsp70 and Hsp100 families were specifically upregulated twelve hours after wounding in root tissue, and exhibited a parallel expression response pattern during recovery from heat stress. Among all Hsf and Hsp families, large expression responses occurred under ultraviolet-B light stress in aerial tissue (shoots) but not subterranean tissue (roots).ConclusionOur findings show that Hsf and Hsp family member genes represent an interaction point between multiple stress response pathways, and therefore warrant functional analysis under conditions apart from heat shock treatment. In addition, our analysis revealed several family and tissue-specific heat shock gene expression patterns that have not been previously described. These results have implications regarding the molecular basis of cross-tolerance in plant species, and raise new questions to be pursued in future experimental studies of the Arabidopsis heat shock response network.

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HEAT SHOCK TRANSCRIPTION FACTOR B2b acts as a transcriptional repressor of VIN3, a gene induced by long-term cold for flowering.

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