Innovation, conservation, and repurposing of gene function in root cell type development.

Plant species have evolved myriads of solutions, including complex cell type development and regulation, to adapt to dynamic environments. To understand this cellular diversity, we profiled tomato root cell type translatomes. Using xylem differentiation in tomato, examples of functional innovation, repurposing, and conservation of transcription factors are described, relative to the model plant Arabidopsis. Repurposing and innovation of genes are further observed within an exodermis regulatory network and illustrate its function. Comparative translatome analyses of rice, tomato, and Arabidopsis cell populations suggest increased expression conservation of root meristems compared with other homologous populations. In addition, the functions of constitutively expressed genes are more conserved than those of cell type/tissue-enriched genes. These observations suggest that higher order properties of cell type and pan-cell type regulation are evolutionarily conserved between plants and animals.

Oligomerization-mediated autoinhibition and cofactor binding of a plant NLR.

Nucleotide-binding leucine-rich repeat (NLR) proteins play a pivotal role in plant immunity by recognizing pathogen effectors1,2. Maintaining a balanced immune response is crucial, as excessive NLR expression can lead to unintended autoimmunity3,4. Unlike most NLRs, the plant NLR required for cell death 2 (NRC2) belongs to a small NLR group characterized by constitutively high expression without self-activation5. The mechanisms underlying NRC2 autoinhibition and activation are not yet understood. Here we show that Solanum lycopersicum (tomato) NRC2 (SlNRC2) forms dimers and tetramers and higher-order oligomers at elevated concentrations. Cryo-electron microscopy shows an inactive conformation of SlNRC2 in these oligomers. Dimerization and oligomerization not only stabilize the inactive state but also sequester SlNRC2 from assembling into an active form. Mutations at the dimeric or interdimeric interfaces enhance pathogen-induced cell death and immunity in Nicotiana benthamiana. The cryo-electron microscopy structures unexpectedly show inositol hexakisphosphate (IP6) or pentakisphosphate (IP5) bound to the inner surface of the C-terminal leucine-rich repeat domain of SlNRC2, as confirmed by mass spectrometry. Mutations at the inositol phosphate-binding site impair inositol phosphate binding of SlNRC2 and pathogen-induced SlNRC2-mediated cell death in N. benthamiana. Our study indicates a negative regulatory mechanism of NLR activation and suggests inositol phosphates as cofactors of NRCs.

A gradient of the HD-Zip regulator Woolly regulates multicellular trichome morphogenesis in tomato.

Homeodomain (HD) proteins regulate embryogenesis in animals such as the fruit fly (Drosophila melanogaster), often in a concentration-dependent manner. HD-leucine zipper (Zip) IV family genes are unique to plants and often function in the L1 epidermal cell layer. However, our understanding of the roles of HD-Zip IV family genes in plant morphogenesis is limited. In this study, we investigated the morphogenesis of tomato (Solanum lycopersicum) multicellular trichomes, a type of micro-organ in plants. We found that a gradient of the HD-Zip IV regulator Woolly (Wo) coordinates spatially polarized cell division and cell expansion in multicellular trichomes. Moreover, we identified a TEOSINTE BRANCHED1, CYCLOIDEA, and PROLIFERATING CELL NUCLEAR ANTIGEN BINDING FACTOR (TCP) transcription factor-encoding gene, SlBRANCHED2a (SlBRC2a), as a key downstream target of Wo that regulates the transition from cell division to cell expansion. High levels of Wo promote cell division in apical trichome cells, whereas in basal trichome cells, Wo mediates a negative feedback loop with SlBRC2a that forces basal cells to enter endoreduplication. The restricted high and low activities of Wo pattern the morphogenesis of tomato multicellular trichomes. These findings provide insights into the functions of HD-Zip IV genes during plant morphogenesis.

The Tomato Guanylate-Binding Protein SlGBP1 Enables Fruit Tissue Differentiation by Maintaining Endopolyploid Cells in a Non-Proliferative State.

Cell fate maintenance is an integral part of plant cell differentiation and the production of functional cells, tissues, and organs. Fleshy fruit development is characterized by the accumulation of water and solutes in the enlarging cells of parenchymatous tissues. In tomato (Solanum lycopersicum), this process is associated with endoreduplication in mesocarp cells. The mechanisms that preserve this developmental program, once initiated, remain unknown. We show here that analysis of a previously identified tomato ethyl methanesulfonate-induced mutant that exhibits abnormal mesocarp cell differentiation could help elucidate determinants of fruit cell fate maintenance. We identified and validated the causal locus through mapping-by-sequencing and gene editing, respectively, and performed metabolic, cellular, and transcriptomic analyses of the mutant phenotype. The data indicate that disruption of the SlGBP1 gene, encoding GUANYLATE BINDING PROTEIN1, induces early termination of endoreduplication followed by late divisions of polyploid mesocarp cells, which consequently acquire the characteristics of young proliferative cells. This study reveals a crucial role of plant GBPs in the control of cell cycle genes, and thus, in cell fate maintenance. We propose that SlGBP1 acts as an inhibitor of cell division, a function conserved with the human hGBP-1 protein.

DNA methylation dynamics of sperm cell lineage development in tomato.

During the sexual reproduction of higher plants, DNA methylation and transcription are broadly changed to reshape a microspore into two sperm cells (SCs) and a vegetative cell (VC). However, when and how the DNA methylation of SCs is established remains not fully understood. Here we investigate the DNA methylation (5 mC) dynamics of SC lineage and the VC in tomato using whole-genome bisulfite sequencing. We find the asymmetric division of the microspore gives its two daughter cells differential methylome. Compared with the generative cell (GC), the VC is hypomethylated at CG sites while hypermethylated at CHG and CHH sites, with the majority of differentially methylation regions targeted to transposable elements (TEs). SCs have a nearly identical DNA methylome to the GC, suggesting that the methylation landscape in SCs may be pre-established following the asymmetric division or inherited from the GC. The random forest classifier for predicting gene and TE expression shows that methylation within the gene body is a more powerful predictor for gene expression. Among all tested samples, gene and TE expression in the microspore may be more predictable by DNA methylation. Our results depict an intact DNA methylome landscape of SC lineage in higher plants, and reveal that the impact of DNA methylation on transcription is variant in different cell types.

Meiotic recombination profiling of interspecific hybrid F1 tomato pollen by linked read sequencing.

Genome wide screening of pooled pollen samples from a single interspecific F1 hybrid obtained from a cross between tomato, Solanum lycopersicum and its wild relative, Solanum pimpinellifolium using linked read sequencing of the haploid nuclei, allowed profiling of the crossover (CO) and gene conversion (GC) landscape. We observed a striking overlap between cold regions of CO in the male gametes and our previously established F6 recombinant inbred lines (RILs) population. COs were overrepresented in non-coding regions in the gene promoter and 5'UTR regions of genes. Poly-A/T and AT rich motifs were found enriched in 1 kb promoter regions flanking the CO sites. Non-crossover associated allelic and ectopic GCs were detected in most chromosomes, confirming that besides CO, GC represents also a source for genetic diversity and genome plasticity in tomato. Furthermore, we identified processed break junctions pointing at the involvement of both homology directed and non-homology directed repair pathways, suggesting a recombination machinery in tomato that is more complex than currently anticipated.

A transcriptomic, metabolomic and cellular approach to the physiological adaptation of tomato fruit to high temperature.

High temperatures can negatively influence plant growth and development. Besides yield, the effects of heat stress on fruit quality traits remain poorly characterised. In tomato, insights into how fruits regulate cellular metabolism in response to heat stress could contribute to the development of heat-tolerant varieties, without detrimental effects on quality. In the present study, the changes occurring in wild type tomato fruits after exposure to transient heat stress have been elucidated at the transcriptome, cellular and metabolite level. An impact on fruit quality was evident as nutritional attributes changed in response to heat stress. Fruit carotenogenesis was affected, predominantly at the stage of phytoene formation, although altered desaturation/isomerisation arose during the transient exposure to high temperatures. Plastidial isoprenoid compounds showed subtle alterations in their distribution within chromoplast sub-compartments. Metabolite profiling suggests limited effects on primary/intermediary metabolism but lipid remodelling was evident. The heat-induced molecular signatures included the accumulation of sucrose and triacylglycerols, and a decrease in the degree of membrane lipid unsaturation, which influenced the volatile profile. Collectively, these data provide valuable insights into the underlying biochemical and molecular adaptation of fruit to heat stress and will impact on our ability to develop future climate resilient tomato varieties.

Overexpression of SlMYB75 enhances resistance to Botrytis cinerea and prolongs fruit storage life in tomato.

KEY MESSAGE: SlMYB75 increased the accumulation of JA and improved the scavenging of excess H2O2 to resist B. cinerea. Overexpression of SlMYB75 greatly prolongs tomato fruit storage life. Botrytis cinerea (B. cinerea) is a major threat to the production and storage life of tomato (Solanum lycopersicum) fruit around the world. SlMYB75 is an R2R3MYB transcription factor associated with the biosynthesis of anthocyanidin, but little is known about its function in the resistance of tomato to B. cinerea. In this study, we found that the overexpression of SlMYB75 regulated the accumulation of jasmonic acid (JA) and promoted the JA-mediated signaling pathway to resist B. cinerea infection. Moreover, the activities of peroxidase and superoxide dismutase, which were activated to scavenge hydrogen peroxide produced as a result of the B. cinerea infection, were enhanced in the transgenic tomato plants. Scanning electron microscopy images showed that the wax on the fruit skin surface was significantly decreased in the transgenic tomatoes compared with the wild type. However, SlMYB75 prolonged fruit storage life by both enhancing resistance to B. cinerea and directly downregulating the fruit shelf life-related gene SlFSR. Collectively, this study provides a good candidate gene for breeding high-quality tomatoes with a long storage life and high disease resistance.

Direct analysis of pollen fitness by flow cytometry: implications for pollen response to stress.

Sexual reproduction in flowering plants depends on the fitness of the male gametophyte during fertilization. Because pollen development is highly sensitive to hot and cold temperature extremes, reliable methods to evaluate pollen viability are important for research into improving reproductive heat stress (HS) tolerance. Here, we describe an approach to rapidly evaluate pollen viability using a reactive oxygen species (ROS) probe dichlorodihydrofluorescein diacetate (i.e. H2 DCFDA-staining) coupled with flow cytometry. In using flow cytometry to analyze mature pollen harvested from Arabidopsis and tomato flowers, we discovered that pollen distributed bimodally into 'low-ROS' and 'high-ROS' subpopulations. Pollen germination assays following fluorescence-activated cell sorting revealed that the high-ROS pollen germinated with a frequency that was 35-fold higher than the low-ROS pollen, supporting a model in which a significant fraction of a flower's pollen remains in a low metabolic or dormant state even after hydration. The ability to use flow cytometry to quantify ROS dynamics within a large pollen population was shown by dose-dependent alterations in DCF-fluorescence in response to oxidative stress or antioxidant treatments. HS treatments (35°C) increased ROS levels, which correlated with a ~60% reduction in pollen germination. These results demonstrate the potential of using flow cytometry-based approaches to investigate metabolic changes during stress responses in pollen.

Optimised production of an anti-fungal antibody in Solanaceae hairy roots to develop new formulations against Candida albicans.

BACKGROUND: Infections caused by fungi are often refractory to conventional therapies and urgently require the development of novel options, such as immunotherapy. To produce therapeutic antibodies, a plant-based expression platform is an attractive biotechnological strategy compared to mammalian cell cultures. In addition to whole plants, hairy roots (HR) cultures can be used, representing an expression system easy to build up, with indefinite growth while handled under containment conditions. RESULTS: In this study the production in HR of a recombinant antibody, proved to be a good candidate for human immunotherapy against fungal infections, is reported. Expression and secretion of this antibody, in an engineered single chain (scFvFc) format, by HR from Nicotiana benthamiana and Solanum lycopersicum have been evaluated with the aim of directly using the deriving extract or culture medium against pathogenic fungi. Although both Solanaceae HR showed good expression levels (up to 68 mg/kg), an optimization of rhizosecretion was only obtained for N. benthamiana HR. A preliminary assessment to explain this result highlighted the fact that not only the presence of proteases, but also the chemical characteristics of the growth medium, can influence antibody yield, with implications on recombinant protein production in HR. Finally, the antifungal activity of scFvFc 2G8 antibody produced in N. benthamiana HR was evaluated in Candida albicans growth inhibition assays, evidencing encouraging results. CONCLUSIONS: Production of this anti-fungal antibody in HR of N. benthamiana and S. lycopersicum elucidated factors affecting pharming in this system and allowed to obtain promising ready-to-use immunotherapeutics against C. albicans.

BZR1 Mediates Brassinosteroid-Induced Autophagy and Nitrogen Starvation in Tomato.

Autophagy, an innate cellular destructive mechanism, plays crucial roles in plant development and responses to stress. Autophagy is known to be stimulated or suppressed by multiple molecular processes, but the role of phytohormone signaling in autophagy is unclear. Here, we demonstrate that the transcripts of autophagy-related genes (ATGs) and the formation of autophagosomes are triggered by enhanced levels of brassinosteroid (BR). Furthermore, the BR-activated transcription factor brassinazole-resistant1 (BZR1), a positive regulator of the BR signaling pathway, is involved in BR-induced autophagy. Treatment with BR enhanced the formation of autophagosomes and the transcripts of ATGs in BZR1-overexpressing plants, while the effects of BR were compromised in BZR1-silenced plants. Yeast one-hybrid analysis and chromatin immunoprecipitation coupled with quantitative polymerase chain reaction revealed that BZR1 bound to the promoters of ATG2 and ATG6 The BR-induced formation of autophagosomes decreased in ATG2- and ATG6-silenced plants. Moreover, exogenous application of BR enhanced chlorophyll content and autophagosome formation and decreased the accumulation of ubiquitinated proteins under nitrogen starvation. Leaf chlorosis and chlorophyll degradation were exacerbated in BZR1-silenced plants and the BR biosynthetic mutant d^im but were alleviated in BZR1- and BZR1-1D-overexpressing plants under nitrogen starvation. Meanwhile, nitrogen starvation-induced expression of ATGs and autophagosome formation were compromised in both BZR1-silenced and d^im plants but were increased in BZR1- and BZR1-1D-overexpressing plants. Taken together, our results suggest that BZR1-dependent BR signaling up-regulates the expression of ATGs and autophagosome formation, which plays a critical role in the plant response to nitrogen starvation in tomato (Solanum lycopersicum).

Coordination of Meristem Doming and the Floral Transition by Late Termination, a Kelch Repeat Protein.

Enlargement and doming of the shoot apical meristem (SAM) is a hallmark of the transition from vegetative growth to flowering. While this change is widespread, its role in the flowering process is unknown. The late termination (ltm) tomato (Solanum lycopersicum) mutant shows severely delayed flowering and precocious doming of the vegetative SAM LTM encodes a kelch domain-containing protein, with no link to known meristem maintenance or flowering time pathways. LTM interacts with the TOPLESS corepressor and with several transcription factors that can provide specificity for its functions. A subgroup of flowering-associated genes is precociously upregulated in vegetative stages of ltm SAMs, among them, the antiflorigen gene SELF PRUNING (SP). A mutation in SP restored the structure of vegetative SAMs in ltm sp double mutants, and late flowering was partially suppressed, suggesting that LTM functions to suppress SP in the vegetative SAM In agreement, SP-overexpressing wild-type plants exhibited precocious doming of vegetative SAMs combined with late flowering, as found in ltm plants. Strong flowering signals can result in termination of the SAM, usually by its differentiation into a flower. We propose that activation of a floral antagonist that promotes SAM growth in concert with floral transition protects it from such terminating effects.

Direct and Indirect Visualization of Bacterial Effector Delivery into Diverse Plant Cell Types during Infection.

To cause disease, diverse pathogens deliver effector proteins into host cells. Pathogen effectors can inhibit defense responses, alter host physiology, and represent important cellular probes to investigate plant biology. However, effector function and localization have primarily been investigated after overexpression in planta. Visualizing effector delivery during infection is challenging due to the plant cell wall, autofluorescence, and low effector abundance. Here, we used a GFP strand system to directly visualize bacterial effectors delivered into plant cells through the type III secretion system. GFP is a beta barrel that can be divided into 11 strands. We generated transgenic Arabidopsis thaliana plants expressing GFP1-10 (strands 1 to 10). Multiple bacterial effectors tagged with the complementary strand 11 epitope retained their biological function in Arabidopsis and tomato (Solanum lycopersicum). Infection of plants expressing GFP1-10 with bacteria delivering GFP11-tagged effectors enabled direct effector detection in planta. We investigated the temporal and spatial delivery of GFP11-tagged effectors during infection with the foliar pathogen Pseudomonas syringae and the vascular pathogen Ralstonia solanacearum Thus, the GFP strand system can be broadly used to investigate effector biology in planta.

Possible Role of Crystal-Bearing Cells in Tomato Fertility and Formation of Seedless Fruits.

Crystal-bearing cells or idioblasts, which deposit calcium oxalate, are located in various tissues and organs of many plant species. The functional significance of their formation is currently unclear. Idioblasts in the leaf parenchyma and the development of crystal-bearing cells in the anther tissues of transgenic tomato plants (Solanum lycopersicon L.), expressing the heterologous FeSOD gene and which showed a decrease in fertility, were studied by transmission and scanning electron microscopy. The amount of calcium oxalate crystals was found to increase significantly in the transgenic plants compared to the wild type (WT) ones in idioblasts and crystal-bearing cells of the upper part of the anther. At the same time, changes in the size and shape of the crystals and their location in anther organs were noted. It seems that the interruption in the break of the anther stomium in transgenic plants was associated with the formation and cell death regulation of a specialized group of crystal-bearing cells. This disturbance caused an increase in the pool of these cells and their localization in the upper part of the anther, where rupture is initiated. Perturbations were also noted in the lower part of the anther in transgenic plants, where the amount of calcium oxalate crystals in crystal-bearing cells was reduced that was accompanied by disturbances in the morphology of pollen grains. Thus, the induction of the formation of crystal-bearing cells and calcium oxalate crystals can have multidirectional effects, contributing to the regulation of oxalate metabolism in the generative and vegetative organs and preventing fertility when the ROS balance changes, in particular, during oxidative stresses accompanying most abiotic and biotic environmental factors.

Organ-wide and ploidy-dependent regulation both contribute to cell-size determination: evidence from a computational model of tomato fruit.

The development of a new organ is the result of coordinated events of cell division and expansion, in strong interaction with each other. This study presents a dynamic model of tomato fruit development that includes cell division, endoreduplication, and expansion processes. The model is used to investigate the potential interactions among these developmental processes within the context of the neo-cellular theory. In particular, different control schemes (either cell-autonomous or organ-controlled) are tested and compared to experimental data from two contrasting genotypes. The model shows that a pure cell-autonomous control fails to reproduce the observed cell-size distribution, and that an organ-wide control is required in order to get realistic cell-size variations. The model also supports the role of endoreduplication as an important determinant of the final cell size and suggests that a direct effect of endoreduplication on cell expansion is needed in order to obtain a significant correlation between size and ploidy, as observed in real data.

Discrete element modelling of tomato tissue deformation and failure at the cellular scale.

Bruise damage in fruit results from cell wall failure and inter-cellular separation. Despite the importance of the micro-mechanics of plant tissue with respect to its integrity, it remains largely unquantified and poorly understood, due to many difficulties during experimental characterization. In this article, a 3D micro-mechanical plant tissue model that is able to model cell rupture and inter-cellular debonding and thus provide more insight into the micro-mechanics was developed. The model is based on the discrete element method (DEM) and represents the tissue as a mass-spring system. Each plant cell is represented as a deformable visco-elastoplastic triangulated mesh under turgor pressure. To model cell wall rupture, it is assumed that a spring connection in the wall breaks at a certain critical stretch ratio and that a ruptured cell is turgorless. The inter-cellular contact model assumes brittle fracture between a cell's node and an adjacent cell's triangle when their bond distance exceeds a critical value. A high-speed tomato fruit cell compression test was simulated and the modelled force-strain curve compares well with the experimental data, including for strains above the elastic limit. By varying the shape of the cell in the compression simulation it was shown that the force-strain curve is highly dependent on the cell shape and thus parameter fitting procedures based on a spherical cell model will be inaccurate. Furthermore, the wall stiffness and thickness showed a positive linear relationship with the force at cell bursting. Besides simulating compression tests of single cells, we also simulated tensile and compression tests on small tissue specimens. Realistic tissue structures of tomato mesocarp tissue were generated by a novel method using DEM simulations of deformable cells in a shrinking cylinder. The cell area, volume and anisotropy distributions of the virtual tissue compared well with micro-CT images of real tomato mesocarp tissue (normalized root mean square error values smaller than 3%). The tissue compression and tensile test simulations demonstrated an important influence of the inter-cellular bonding energy and tissue porosity on the tissue failure characteristics and elastic modulus.

Yellow-emissive carbon dots with a large Stokes shift are viable fluorescent probes for detection and cellular imaging of silver ions and glutathione.

Yellow-emissive carbon dots (Y-CDs) were prepared by a solvothermal method using anhydrous citric acid and 2,3-phenazinediamine as the starting materials. The Y-CDs display a 24% fluorescence quantum yield, a 188-nm Stokes' shift and excellent stability. They are shown here to be excellent fluorescent probes for the determination of Ag(I) ion and glutathione (GSH). If exposed to Ag(I) ions, they are bound by the carboxy groups of the Y-CDs, and this causes quenching of fluorescence (with excitation/emission maxima at 380/568 nm) via a static quenching mechanism. This effect was used to design a fluorometric assay for Ag(I). The quenched fluorescence of the Y-CDs can be restored by adding GSH due to the high affinity of GSH for Ag(I). The calibration plot for Ag(I) is linear in the 1-4 µM Ag(I) concentration range, and the limit of detection is 31 nM. The respective values for GSH are 5-32 µM, and 76 nM, respectively. The method was applied to the detection of Ag(I) in spiked environmental water samples and gave recoveries ranging from 93 to 107%. It was also applied to the determination of GSH in tomatoes and purple grapes and gave satisfactory recoveries. The Y-CDs display low cytotoxicity and were successfully used to image Ag(I) and GSH in H1299 cells. Graphical abstract Schematic presentation of the mechanism of yellow fluorescent CDs for the detection of Ag+ and glutathione.

Stem cell activation by light guides plant organogenesis.

Leaves originate from stem cells located at the shoot apical meristem. The meristem is shielded from the environment by older leaves, and leaf initiation is considered to be an autonomous process that does not depend on environmental cues. Here we show that light acts as a morphogenic signal that controls leaf initiation and stabilizes leaf positioning. Leaf initiation in tomato shoot apices ceases in the dark but resumes in the light, an effect that is mediated through the plant hormone cytokinin. Dark treatment also affects the subcellular localization of the auxin transporter PIN1 and the concomitant formation of auxin maxima. We propose that cytokinin is required for meristem propagation, and that auxin redirects cytokinin-inducible meristem growth toward organ formation. In contrast to common wisdom over the last 150 years, the light environment controls the initiation of lateral organs by regulating two key hormones: auxin and cytokinin.

S-Nitrosylation inhibits the kinase activity of tomato phosphoinositide-dependent kinase 1 (PDK1).

It is well known that the reactive oxygen species NO can trigger cell death in plants and other organisms, but the underlying molecular mechanisms are not well understood. Here we provide evidence that NO may trigger cell death in tomato (Solanum lycopersicum) by inhibiting the activity of phosphoinositide-dependent kinase 1 (SlPDK1), a conserved negative regulator of cell death in yeasts, mammals, and plants, via S-nitrosylation. Biotin-switch assays indicated that SlPDK1 is a target of S-nitrosylation. Moreover, the kinase activity of SlPDK1 was inhibited by S-nitrosoglutathione in a concentration-dependent manner, indicating that SlPDK1 activity is abrogated by S-nitrosylation. The S-nitrosoglutathione-induced inhibition was reversible in the presence of a reducing agent but additively enhanced by hydrogen peroxide (H2O2). Our LC-MS/MS analyses further indicated that SlPDK1 is primarily S-nitrosylated on a cysteine residue at position 128 (Cys128), and substitution of Cys128 with serine completely abolished SlPDK1 kinase activity, suggesting that S-nitrosylation of Cys128 is responsible for SlPDK1 inhibition. In summary, our results establish a potential link between NO-triggered cell death and inhibition of the kinase activity of tomato PDK1.

DNA-binding and repressor function are prerequisites for the turnover of the tomato heat stress transcription factor HsfB1.

HsfB1 is a central regulator of heat stress (HS) response and functions dually as a transcriptional co-activator of HsfA1a and a general repressor in tomato. HsfB1 is efficiently synthesized during the onset of HS and rapidly removed in the course of attenuation during the recovery phase. Initial results point to a complex regime modulating HsfB1 abundance involving the molecular chaperone Hsp90. However, the molecular determinants affecting HsfB1 stability needed to be established. We provide experimental evidence that DNA-bound HsfB1 is efficiently targeted for degradation when active as a transcriptional repressor. Manipulation of the DNA-binding affinity by mutating the HsfB1 DNA-binding domain directly influences the stability of the transcription factor. During HS, HsfB1 is stabilized, probably due to co-activator complex formation with HsfA1a. The process of HsfB1 degradation involves nuclear localized Hsp90. The molecular determinants of HsfB1 turnover identified in here are so far seemingly unique. A mutational switch of the R/KLFGV repressor motif's arginine and lysine implies that the abundance of other R/KLFGV type Hsfs, if not other transcription factors as well, might be modulated by a comparable mechanism. Thus, we propose a versatile mechanism for strict abundance control of the stress-induced transcription factor HsfB1 for the recovery phase, and this mechanism constitutes a form of transcription factor removal from promoters by degradation inside the nucleus.