Promoter specificity and interactions between early and late Arabidopsis heat shock factors.
The class A heat shock factors HsfA1a and HsfA1b are highly conserved, interacting regulators, responsible for the immediate-early transcription of a subset of heat shock genes in Arabidopsis. In order to determine functional cooperation between them, we used a reporter assay based on transient over-expression in Arabidopsis protoplasts. Reporter plasmids containing promoters of Hsf target genes fused with the GFP coding region were co-transformed with Hsf effector plasmids. The GFP reporter gene activity was quantified using flow cytometry. Three of the tested target gene promoters (Hsp25.3, Hsp18.1-CI, Hsp26.5) resulted in a strong reporter gene activity, with HsfA1a or HsfA1b alone, and significantly enhanced GFP fluorescence when both effectors were co-transformed. A second set of heat shock promoters (HsfA2, Hsp17.6CII, Hsp17.6C-CI) was activated to much lower levels. These data suggest that HsfA1a/1b cooperate synergistically at a number of target gene promoters. These targets are also regulated via the late HsfA2, which is the most strongly heat-induced class A-Hsf in Arabidopsis. HsfA2 has also the capacity to interact with HsfA1a and HsfA1b as determined by bimolecular fluorescence complementation (BiFC) in Arabidopsis protoplasts and yeast-two-hybrid assay. However, there was no synergistic effect on Hsp18.1-CI promoter-GFP reporter gene expression when HsfA2 was co-expressed with either HsfA1a or HsfA1b. These data provide evidence that interaction between early and late HSF is possible, but only interaction between the early Hsfs results in a synergistic enhancement of expression of certain target genes. The interaction of HsfA1a/A1b with the major-late HsfA2 may possibly support recruitment of HsfA2 and replacement of HsfA1a/A1b at the same target gene promoters.
Li
Ming
M
Zentrum für Molekularbiologie der Pflanzen (ZMBP), Allgemeine Genetik, Universität Tübingen, Auf der Morgenstelle 28, 72076 Tübingen, Germany.
Berendzen
Kenneth W
KW
Schöffl
Friedrich
F
D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D021381:Protein Transport; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D029681:Arabidopsis Proteins; D004268:DNA-Binding Proteins; C000620134:HSFA2 protein, Arabidopsis; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins; D010940:Plant Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins
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