BZR1 and BES1 transcription factors mediate brassinosteroid control over root system architecture in response to nitrogen availability.

Plants modify their root system architecture (RSA) in response to nitrogen (N) deficiency. The plant steroidal hormone, brassinosteroid (BR), plays important roles in root growth and development. This study demonstrates that optimal levels of exogenous BR impact significant increases in lateral root length and numbers in Arabidopsis seedlings under mild N-deficient conditions as compared to untreated seedlings. The impact of BR on RSA was stronger under mild N deficiency than under N-sufficient conditions. The BR effects on RSA were mimicked in dominant mutants of BZR1 and BES1 (bzr1-1D and bes1-D) transcription factors, while the RSA was highly reduced in the BR-insensitive mutant bri1-6, confirming that BR signaling is essential for the development of RSA under both N-sufficient and N-deficient conditions. Exogenous BR and constitutive activity of BZR1 and BES1 in dominant mutants led to enhanced root meristem, meristematic cell number, and cortical cell length. Under mild N deficiency, bzr1-1D displayed higher fresh and dry shoot weights, chlorophyll content, and N levels in the shoot, as compared to the wild type. These results indicate that BR modulates RSA under both N-sufficient and N-deficient conditions via the transcription factors BES1/BZR1 module and confers tolerance to N deficiency.

Improving Blueberry Fruit Nutritional Quality through Physiological and Genetic Interventions: A Review of Current Research and Future Directions.

Blueberry, hailed as an antioxidant superfood, is the fruit of small shrubs in the genus Vaccinium (family Ericaceae). The fruits are a rich source of vitamins, minerals and antioxidants such as flavonoids and phenolic acids. The antioxidative and anti-inflammatory activities derived from the polyphenolic compounds, particularly from the abundantly present anthocyanin pigment, have been highlighted as the major contributing factor to the health-benefitting properties of blueberry. In recent years, blueberry cultivation under polytunnels has expanded, with plastic covers designed to offer protection of crop and fruit yield from suboptimal environmental conditions and birds. An important consideration is that the covers reduce photosynthetically active radiation (PAR) and filter out ultraviolet (UV) radiation that is critical for the fruit's bioactive composition. Blueberry fruits grown under covers have been reported to have reduced antioxidant capacity as compared to fruits from open fields. In addition to light, abiotic stresses such as salinity, water deficit, and low temperature trigger accumulation of antioxidants. We highlight in this review how interventions such as light-emitting diodes (LEDs), photo-selective films, and exposure of plants to mild stresses, alongside developing new varieties with desired traits, could be used to optimise the nutritional quality, particularly the content of polyphenols, of blueberry grown under covers.

Tangerine tomato roots show increased accumulation of acyclic carotenoids, less abscisic acid, drought sensitivity, and impaired endomycorrhizal colonization.

The Heirloom Golden tangerine tomato fruit variety is highly nutritious due to accumulation of tetra-cis-lycopene, that has a higher bioavailability and recognised health benefits in treating anti-inflammatory diseases compared to all-trans-lycopene isomers found in red tomatoes. We investigated if photoisomerization of tetra-cis-lycopene occurs in roots of the MicroTom tangerine (tangmic) tomato and how this affects root to shoot biomass, mycorrhizal colonization, abscisic acid accumulation, and responses to drought. tangmic plants grown in soil under glasshouse conditions displayed a reduction in height, number of flowers, fruit yield, and root length compared to wild-type (WT). Soil inoculation with Rhizophagus irregularis revealed fewer arbuscules and other fungal structures in the endodermal cells of roots in tangmic relative to WT. The roots of tangmic hyperaccumulated acyclic cis-carotenes, while only trace levels of xanthophylls and abscisic acid were detected. In response to a water deficit, leaves from the tangmic plants displayed a rapid decline in maximum quantum yield of photosystem II compared to WT, indicating a defective root to shoot signalling response to drought. The lack of xanthophylls biosynthesis in tangmic roots reduced abscisic acid levels, thereby likely impairing endomycorrhizal colonisation and drought-induced root to shoot signalling.

Identification of microRNAs involved in lipid biosynthesis and seed size in developing sea buckthorn seeds using high-throughput sequencing.

Sea buckthorn is a plant of medicinal and nutritional importance owing in part to the high levels of essential fatty acids, linoleic (up to 42%) and α-linolenic (up to 39%) acids in the seed oil. Sea buckthorn can produce seeds either via the sexual pathway or by apomixis. The seed development and maturation programs are critically dependent on miRNAs. To understand miRNA-mediated regulation of sea buckthorn seed development, eight small RNA libraries were constructed for deep sequencing from developing seeds of a low oil content line 'SJ1' and a high oil content line 'XE3'. High-throughput sequencing identified 137 known miRNA from 27 families and 264 novel miRNAs. The potential targets of the identified miRNAs were predicted based on sequence homology. Nineteen (four known and 15 novel) and 22 (six known and 16 novel) miRNAs were found to be involved in lipid biosynthesis and seed size, respectively. An integrated analysis of mRNA and miRNA transcriptome and qRT-PCR identified some key miRNAs and their targets (miR164d-ARF2, miR168b-Δ9D, novelmiRNA-108-ACC, novelmiRNA-23-GPD1, novelmiRNA-58-DGAT1, and novelmiRNA-191-DGAT2) potentially involved in seed size and lipid biosynthesis of sea buckthorn seed. These results indicate the potential importance of miRNAs in regulating lipid biosynthesis and seed size in sea buckthorn.

Brassinosteroid Action in Plant Abiotic Stress Tolerance.

Brassinosteroids (BRs) are a class of plant steroidal hormones that play essential roles in plant growth and development. Systematic studies had first been undertaken concomitantly to determine both the effects of exogenous BR on stress phenotypes of Arabidopsis thaliana and Brassica napus (rapeseed) seedlings and the expression of stress marker genes in BR-treated and untreated seedlings. When reproducible and convincing evidence of the role of BR in stress tolerance had been obtained, molecular mechanisms underlying the ability of BR to confer tolerance against heat, cold, drought, and salt stress, as well as pathogen resistance were studied with several molecular approaches and tools. The results of these studies have together provided valuable insights into how BRs, through their control of many basic cellular processes and stress responses, promote vigor in plants and prepare the plant to mount a dynamic response upon environmental challenges. Protocols to assess BR effects on abiotic stress tolerance in Arabidopsis and rapeseed seedlings are described here and they can be fine-tuned and adapted for other plant species.

Overexpression of the brassinosteroid biosynthetic gene DWF4 in Brassica napus simultaneously increases seed yield and stress tolerance.

As a resource allocation strategy, plant growth and defense responses are generally mutually antagonistic. Brassinosteroid (BR) regulates many aspects of plant development and stress responses, however, genetic evidence of its integrated effects on plant growth and stress tolerance is lacking. We overexpressed the Arabidopsis BR biosynthetic gene AtDWF4 in the oilseed plant Brassica napus and scored growth and stress response phenotypes. The transgenic B. napus plants, in comparison to wild type, displayed increased seed yield leading to increased overall oil content per plant, higher root biomass and root length, significantly better tolerance to dehydration and heat stress, and enhanced resistance to necrotrophic fungal pathogens Leptosphaeria maculans and Sclerotinia sclerotiorum. Transcriptome analysis supported the integrated effects of BR on growth and stress responses; in addition to BR responses associated with growth, a predominant plant defense signature, likely mediated by BES1/BZR1, was evident in the transgenic plants. These results establish that BR can interactively and simultaneously enhance abiotic and biotic stress tolerance and plant productivity. The ability to confer pleiotropic beneficial effects that are associated with different agronomic traits suggests that BR-related genes may be important targets for simultaneously increasing plant productivity and performance under stress conditions.

Gene expression and functional analyses in brassinosteroid-mediated stress tolerance.

The plant hormone brassinosteroid (BR) plays essential roles in plant growth and development, while also controlling plant stress responses. This dual ability of BR is intriguing from a mechanistic point of view and as a viable solution for stabilizing crop yields under the changing climatic conditions. Here we report a time course analysis of BR responses under both stress and no-stress conditions, the results of which establish that BR incorporates many stress-related features even under no-stress conditions, which are then accompanied by a dynamic stress response under unfavourable conditions. Found within the BR transcriptome were distinct molecular signatures of two stress hormones, abscisic acid and jasmonic acid, which were correlated with enhanced endogenous levels of the two hormones in BR-treated seedlings. The marked presence of genes related to protein metabolism and modification, defence responses and calcium signalling highlights the significance of their associated mechanisms and roles in BR processes. Functional analysis of loss-of-function mutants of a subset of genes selected from the BR transcriptome identified abiotic stress-related roles for ACID PHOSPHATASE5 (ACP5), WRKY33, JACALIN-RELATED LECTIN1-3 (JAC-LEC1-3) and a BR-RESPONSIVE-RECEPTOR-LIKE KINASE (BRRLK). Overall, the results of this study provide a clear link between the molecular changes impacted by BR and its ability to confer broad-range stress tolerance, emphasize the importance of post-translational modification and protein turnover as BR regulatory mechanisms and demonstrate the BR transcriptome as a repertoire of new stress-related regulatory and structural genes.

Metabolite profiling and expression analysis of flavonoid, vitamin C and tocopherol biosynthesis genes in the antioxidant-rich sea buckthorn (Hippophae rhamnoides L.).

In this study, phenolic compounds were analyzed in developing berries of four Canadian grown sea buckthorn (Hippophae rhamnoides L.) cultivars ('RC-4', 'E6590', 'Chuyskaya' and 'Golden Rain') and in leaves of two of these cultivars. Among phenolic acids, p-coumaric acid was the highest in berries, while gallic acid was predominant in leaves. In the flavonoid class of compounds, myricetin/rutin, kaempferol, quercetin and isorhamnetin were detected in berries and leaves. Berries of the 'RC-4' cultivar had approximately ⩾ 2-fold higher levels of myricetin and quercetin at 17.5mg and 17.2 mg/100 g FW, respectively, than the other cultivars. The flavonoid content in leaves was considerably more than in berries with rutin and quercetin levels up to 135 mg and 105 mg/100 g FW, respectively. Orthologs of 15 flavonoid biosynthesis pathway genes were identified within the transcriptome of sea buckthorn mature seeds. Semi-quantitative RT-PCR analysis of these genes in developing berries indicated relatively higher expression of genes such as CHS, F3'H, DFR and LDOX in the 'RC-4' cultivar than in the 'Chuyskaya' cultivar. Vitamin C levels in ripened berries of the Canadian cultivars were on the high end of the concentration range reported for most other sea buckthorn cultivars. Orthologs of genes involved in vitamins C and E biosynthesis were also identified, expanding the genomic resources for this nutritionally important plant.

Reciprocal responses in the interaction between Arabidopsis and the cell-content-feeding chelicerate herbivore spider mite.

Most molecular-genetic studies of plant defense responses to arthropod herbivores have focused on insects. However, plant-feeding mites are also pests of diverse plants, and mites induce different patterns of damage to plant tissues than do well-studied insects (e.g. lepidopteran larvae or aphids). The two-spotted spider mite (Tetranychus urticae) is among the most significant mite pests in agriculture, feeding on a staggering number of plant hosts. To understand the interactions between spider mite and a plant at the molecular level, we examined reciprocal genome-wide responses of mites and its host Arabidopsis (Arabidopsis thaliana). Despite differences in feeding guilds, we found that transcriptional responses of Arabidopsis to mite herbivory resembled those observed for lepidopteran herbivores. Mutant analysis of induced plant defense pathways showed functionally that only a subset of induced programs, including jasmonic acid signaling and biosynthesis of indole glucosinolates, are central to Arabidopsis's defense to mite herbivory. On the herbivore side, indole glucosinolates dramatically increased mite mortality and development times. We identified an indole glucosinolate dose-dependent increase in the number of differentially expressed mite genes belonging to pathways associated with detoxification of xenobiotics. This demonstrates that spider mite is sensitive to Arabidopsis defenses that have also been associated with the deterrence of insect herbivores that are very distantly related to chelicerates. Our findings provide molecular insights into the nature of, and response to, herbivory for a representative of a major class of arthropod herbivores.

Fatty acid composition of developing sea buckthorn (Hippophae rhamnoides L.) berry and the transcriptome of the mature seed.

BACKGROUND: Sea buckthorn (Hippophae rhamnoides L.) is a hardy, fruit-producing plant known historically for its medicinal and nutraceutical properties. The most recognized product of sea buckthorn is its fruit oil, composed of seed oil that is rich in essential fatty acids, linoleic (18:2 ω-6) and α-linolenic (18:3 ω-3) acids, and pulp oil that contains high levels of monounsaturated palmitoleic acid (16:1 ω-7). Sea buckthorn is fast gaining popularity as a source of functional food and nutraceuticals, but currently has few genomic resources; therefore, we explored the fatty acid composition of Canadian-grown cultivars (ssp. mongolica) and the sea buckthorn seed transcriptome using the 454 GS FLX sequencing technology. RESULTS: GC-MS profiling of fatty acids in seeds and pulp of berries indicated that the seed oil contained linoleic and α-linolenic acids at 33-36% and 30-36%, respectively, while the pulp oil contained palmitoleic acid at 32-42%. 454 sequencing of sea buckthorn cDNA collections from mature seeds yielded 500,392 sequence reads, which identified 89,141 putative unigenes represented by 37,482 contigs and 51,659 singletons. Functional annotation by Gene Ontology and computational prediction of metabolic pathways indicated that primary metabolism (protein>nucleic acid>carbohydrate>lipid) and fatty acid and lipid biosynthesis pathways were highly represented categories. Sea buckthorn sequences related to fatty acid biosynthesis genes in Arabidopsis were identified, and a subset of these was examined for transcript expression at four developing stages of the berry. CONCLUSION: This study provides the first comprehensive genomic resources represented by expressed sequences for sea buckthorn, and demonstrates that the seed oil of Canadian-grown sea buckthorn cultivars contains high levels of linoleic acid and α-linolenic acid in a close to 1:1 ratio, which is beneficial for human health. These data provide the foundation for further studies on sea buckthorn oil, the enzymes involved in its biosynthesis, and the genes involved in the general hardiness of sea buckthorn against environmental conditions.

Brassinosteroid leaf unrolling QTL mapping in durum wheat.

Brassinosteroids are a newly reported class of plant growth phytohormones found in plants throughout the plant kingdom. Functioning at very low concentrations, they play an essential role in improving biomass yield and stress tolerance. There are no reports in the literature of the genetic variability of responsiveness of brassinosteroids in wheat; most studies on brassinosteroids have focused on the physiological effects of exogenous addition of brassinosteroids. Our aim was to study the genetic variation in the responsiveness of a doubled haploid durum wheat population to three brassinosteroid concentrations using the leaf unrolling test, which is a simple bioassay to test brassinosteroid activity. An F(1)-derived doubled haploid population of 77 individuals from the cross Strongfield/Blackbird was used to construct a genetic map of 427 molecular marker loci. The leaf unrolling test was performed on the parents and doubled haploid genotypes of the population using 0.2, 2 and 20 nM brassinosteroid concentrations. The results indicated significant differences in leaf unrolling between the two parents, doubled haploid genotypes, treatments and genotype-by-treatment combinations. Transgressive segregation beyond Strongfield of leaf unrolling was observed for all concentrations, with the strongest response at 20 nM. Putative quantitative trait loci were revealed in the intervals Xgwm2-Xbarc45 on chromosome 3A and Xwmc643a-Xwmc625a on chromosome 3B. Additional quantitative trait loci were associated with markers Xwmc48a, Xwmc511, Xwmc89a and Xgwmc692 on chromosome 4B, and Xwmc17 on chromosome 7A. This work should enhance the understanding of the relationship between stress tolerance and productivity, and responsiveness to brassinosteroids.

In silico identification of carboxylate clamp type tetratricopeptide repeat proteins in Arabidopsis and rice as putative co-chaperones of Hsp90/Hsp70.

The essential eukaryotic molecular chaperone Hsp90 operates with the help of different co-chaperones, which regulate its ATPase activity and serve as adaptors to recruit client proteins and other molecular chaperones, such as Hsp70, to the Hsp90 complex. Several Hsp90 and Hsp70 co-chaperones contain the tetratricopeptide repeat (TPR) domain, which interacts with the highly conserved EEVD motif at the C-terminal ends of Hsp90 and Hsp70. The acidic side chains in EEVD interact with a subset of basic residues in the TPR binding pocket called a 'carboxylate clamp'. Since the carboxylate clamp residues are conserved in the TPR domains of known Hsp90/Hsp70 co-chaperones, we carried out an in silico search for TPR proteins in Arabidopsis and rice comprising of at least one three-motif TPR domain with conserved amino acid residues required for Hsp90/Hsp70 binding. This approach identified in Arabidopsis a total of 36 carboxylate clamp (CC)-TPR proteins, including 24 novel proteins, with potential to interact with Hsp90/Hsp70. The newly identified CC-TPR proteins in Arabidopsis and rice contain additional protein domains such as ankyrin, SET, octicosapeptide/Phox/Bem1p (Phox/PB1), DnaJ-like, thioredoxin, FBD and F-box, and protein kinase and U-box, indicating varied functions for these proteins. To provide proof-of-concept of the newly identified CC-TPR proteins for interaction with Hsp90, we demonstrated interaction of AtTPR1 and AtTPR2 with AtHsp90 in yeast two-hybrid and in vitro pull down assays. These findings indicate that the in silico approach used here successfully identified in a genome-wide context CC-TPR proteins with potential to interact with Hsp90/Hsp70, and further suggest that the Hsp90/Hsp70 system relies on TPR co-chaperones more than it was previously realized.

Brassinosteroid-mediated stress tolerance in Arabidopsis shows interactions with abscisic acid, ethylene and salicylic acid pathways.

BACKGROUND: Brassinosteroids (BRs) play crucial roles in plant development and also promote tolerance to a range of abiotic stresses. Although much has been learned about their roles in plant development, the mechanisms by which BRs control plant stress responses and regulate stress-responsive gene expression are not fully known. Since BR interacts with other plant hormones, it is likely that the stress tolerance conferring ability of BR lies in part in its interactions with other stress hormones. RESULTS: Using a collection of Arabidopsis mutants that are either deficient in or insensitive to abscisic acid (ABA), ethylene (ET), jasmonic acid (JA) and salicylic acid (SA), we studied the effects of 24-epibrassinloide (EBR) on basic thermotolerance and salt tolerance of these mutants. The positive impact of EBR on thermotolerance in proportion to wild type was evident in all mutants studied, with the exception of the SA-insensitive npr1-1 mutant. EBR could rescue the ET-insensitive ein2 mutant from its hypersensitivity to salt stress-induced inhibition of seed germination, but remained ineffective in increasing the survival of eto1-1 (ET-overproducer) and npr1-1 seedlings on salt. The positive effect of EBR was significantly greater in the ABA-deficient aba1-1 mutant as compared to wild type, indicating that ABA masks BR effects in plant stress responses. Treatment with EBR increased expression of various hormone marker genes in both wild type and mutant seedlings, although to different levels. CONCLUSIONS: These results together indicate that the redox-sensitive protein NPR1 (NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1), a master regulator of SA-mediated defense genes, is likely a critical component of EBR-mediated increase in thermotolerance and salt tolerance, but it is not required for EBR-mediated induction of PR-1 (PATHOGENESIS-RELATED1) gene expression; that BR exerts anti-stress effects independently as well as through interactions with other hormones; that ABA inhibits BR effects during stress; and that BR shares transcriptional targets with other hormones.

Characterization of plant p23-like proteins for their co-chaperone activities.

The small acidic protein p23 is best described as a co-chaperone of Hsp90, an essential molecular chaperone in eukaryotes. p23 binds to the ATP-bound form of Hsp90 and stabilizes the Hsp90-client protein complex by slowing down ATP turnover. The stabilizing activity of p23 was first characterized in studies of steroid receptor-Hsp90 complexes. Earlier studies of the Hsp90 chaperone complex in plants suggested that a p23-like stabilizing activity was absent in plant cell lysates. Here, we show that p23-like proteins are present in plants and are capable of binding Hsp90, but unlike human p23 and yeast ortholog Sba1, the plant p23-like proteins do not stabilize the steroid receptor-Hsp90 complexes formed in wheat germ lysate. Furthermore, these proteins do not inhibit the ATPase activity of plant Hsp90. While transcripts of Arabidopsis thaliana p23-1 and Atp23-2 were detected under normal growing conditions, those of the closely related Brassica napus p23-1 were present only after moderate heat stress. These observations suggest that p23-like proteins in plants are conserved in their binding to Hsp90 but have evolved mechanisms of action different from their yeast and animal counterparts.

The Hop/Sti1-Hsp90 chaperone complex facilitates the maturation and transport of a PAMP receptor in rice innate immunity.

Recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs) represents a critical first step of innate defense in plants and animals. However, maturation and transport of PRRs are not well understood. We find that the rice chitin receptor OsCERK1 interacts with Hsp90 and its cochaperone Hop/Sti1 in the endoplasmic reticulum (ER). Hop/Sti1 and Hsp90 are required for efficient transport of OsCERK1 from the ER to the plasma membrane (PM) via a pathway dependent on Sar1, a small GTPase which regulates ER-to-Golgi trafficking. Further, Hop/Sti1 and Hsp90 are present at the PM in a complex (designated the "defensome") with OsRac1, a plant-specific Rho-type GTPase. Finally, Hop/Sti1 was required for chitin-triggered immunity and resistance to rice blast fungus. Our results suggest that the Hop/Sti1-Hsp90 chaperone complex plays an important and likely conserved role in the maturation and transport of PRRs and may function to link PRRs and Rac/Rop GTPases.

Brassinosteroid: a biotechnological target for enhancing crop yield and stress tolerance.

Brassinosteroids (BRs) are a group of naturally occurring plant steroidal compounds with wide ranging biological activity. Because BRs control several important agronomic traits such as flowering time, plant architecture, seed yield and stress tolerance, the genetic manipulation of BR biosynthesis, conversion or perception offers a unique possibility of significantly increasing crop yields through both changing plant metabolism and protecting plants from environmental stresses. Genetic manipulation of BR activity has indeed led to increases in crop yield by 20-60%, confirming the value of further research on BRs to improve productivity.

Brassinosteroid confers tolerance in Arabidopsis thaliana and Brassica napus to a range of abiotic stresses.

In addition to an essential role in plant development, brassinosteroids (BRs) appear to have the ability to protect plants against various environmental stresses. However, studies confirming the ability of BRs to modulate plant responses to different environmental stresses are lacking. Earlier we had demonstrated that treatment with 24-epibrassinolide (EBR), a BR, increases the basic thermotolerance of Brassica napus and tomato seedlings [Plant Mol Biol 40:333-342, 1999]. Here we demonstrate that EBR treatment enhances seedling tolerance to drought and cold stresses in both Arabidopsis thaliana and B. napus, and helps to overcome a salt-stress-induced inhibition of seed germination. The ability of EBR to confer tolerance in plants to a variety of stresses was confirmed through analysis of expression of a subset of drought and cold stress marker genes. Transcriptional changes in these genes were more apparent in EBR-treated A. thaliana, in particular during earlier time points of stress. To see if BR is essential for the heat stress (HS) response, we made use of BR-deficient mutants. Both det2-1 and dwf4 mutants still expressed heat shock proteins (hsps) to high levels during HS, indicating that although BR augments thermotolerance in plants, it is not necessary for hsp expression during HS.

Brassinosteroid-Mediated Stress Responses.

Brassinosteroids (BRs) are a group of naturally occurring plant steroidal compounds with wide-ranging biological activity that offer the unique possibility of increasing crop yields through both changing plant metabolism and protecting plants from environmental stresses. In recent years, genetic and biochemical studies have established an essential role for BRs in plant development, and on this basis BRs have been given the stature of a phytohormone. A remarkable feature of BRs is their potential to increase resistance in plants to a wide spectrum of stresses, such as low and high temperatures, drought, high salt, and pathogen attack. Despite this, only a few studies aimed at understanding the mechanism by which BRs promote stress resistance have been undertaken. Studies of the BR signaling pathway and BR gene-regulating properties indicate that there is cross-talk between BRs and other hormones, including those with established roles in plant defense responses such as abscisic acid, jasmonic acid, and ethylene. Recent studies aimed at understanding how BRs modulate stress responses suggest that complex molecular changes underlie BR-induced stress tolerance in plants. Analyses of these changes should generate exciting results in the future and clarify whether the ability of BRs to increase plant resistance to a range of stresses lies in the complex interactions of BRs with other hormones. Future studies should also elucidate if BRI1, an essential component of the BR receptor, directly participates in stress response signaling through interactions with ligands and proteins involved in plant defense responses.

Cytosolic HSP90 associates with and modulates the Arabidopsis RPM1 disease resistance protein.

The Arabidopsis protein RPM1 activates disease resistance in response to Pseudomonas syringae proteins targeted to the inside of the host cell via the bacterial type III delivery system. We demonstrate that specific mutations in the ATP-binding domain of a single Arabidopsis cytosolic HSP90 isoform compromise RPM1 function. These mutations do not affect the function of related disease resistance proteins. RPM1 associates with HSP90 in plant cells. The Arabidopsis proteins RAR1 and SGT1 are required for the action of many R proteins, and display some structural similarity to HSP90 co-chaperones. Each associates with HSP90 in plant cells. Our data suggest that (i) RPM1 is an HSP90 client protein; and (ii) RAR1 and SGT1 may function independently as HSP90 cofactors. Dynamic interactions among these proteins can regulate RPM1 stability and function, perhaps similarly to the formation and regulation of animal steroid receptor complexes.

Characterization of a plant homolog of hop, a cochaperone of hsp90.

The 90-kD molecular chaperone hsp90 is the key component of a multiprotein chaperone complex that facilitates folding, stabilization, and functional modulation of a number of signaling proteins. The components of the animal chaperone complex include hsp90, hsp70, hsp40, Hop, and p23. The animal Hop functions to link hsp90 and hsp70, and it can also inhibit the ATPase activity of hsp90. We have demonstrated the presence of an hsp90 chaperone complex in plant cells, but not all components of the complex have been identified. Here, we report the isolation and characterization of soybean (Glycine max) GmHop-1, a soybean homolog of mammalian Hop. An analysis of soybean expressed sequence tags, combined with preexisting data in literature, suggested the presence of at least three related genes encoding Hop-like proteins in soybean. Transcripts corresponding to Hop-like proteins in soybean were detected under normal growth conditions, and their levels increased further in response to stress. A recombinant GmHop-1 bound hsp90 and its binding to hsp90 could be blocked by the tetratricopeptide repeat (TPR) domain of rat (Rattus norvegicus) protein phosphatase 5. Deletion of amino acids 325 to 395, adjacent to the TPR2A domain in GmHop-1, resulted in loss of hsp90 binding. In a minimal assembly system, GmHop-1 was able to stimulate mammalian steroid receptor folding. These data show that plant and animal Hop homologs are conserved in their general characteristics, and suggest that a Hop-like protein in plants is an important cochaperone of plant hsp90.