Varying Atmospheric CO2 Mediates the Cold-Induced CBF-Dependent Signaling Pathway and Freezing Tolerance in Arabidopsis.

Changes in the stomatal aperture in response to CO2 levels allow plants to manage water usage, optimize CO2 uptake and adjust to environmental stimuli. The current study reports that sub-ambient CO2 up-regulated the low temperature induction of the C-repeat Binding Factor (CBF)-dependent cold signaling pathway in Arabidopsis (Arabidopsis thaliana) and the opposite occurred in response to supra-ambient CO2. Accordingly, cold induction of various downstream cold-responsive genes was modified by CO2 treatments and expression changes were either partially or fully CBF-dependent. Changes in electrolyte leakage during freezing tests were correlated with CO2's effects on CBF expression. Cold treatments were also performed on Arabidopsis mutants with altered stomatal responses to CO2, i.e., high leaf temperature 1-2 (ht1-2, CO2 hypersensitive) and ß-carbonic anhydrase 1 and 4 (ca1ca4, CO2 insensitive). The cold-induced expression of CBF and downstream CBF target genes plus freezing tolerance of ht1-2 was consistently less than that for Col-0, suggesting that HT1 is a positive modulator of cold signaling. The ca1ca4 mutant had diminished CBF expression during cold treatment but the downstream expression of cold-responsive genes was either similar to or greater than that of Col-0. This finding suggested that ßCA1/4 modulates the expression of certain cold-responsive genes in a CBF-independent manner. Stomatal conductance measurements demonstrated that low temperatures overrode low CO2-induced stomatal opening and this process was delayed in the cold tolerant mutant, ca1ca4, compared to the cold sensitive mutant, ht1-2. The similar stomatal responses were evident from freezing tolerant line, Ox-CBF, overexpression of CBF3, compared to wild-type ecotype Ws-2. Together, these results indicate that CO2 signaling in stomata and CBF-mediated cold signaling work coordinately in Arabidopsis to manage abiotic stress.

Ethylene and RIPENING INHIBITOR Modulate Expression of SlHSP17.7A, B Class I Small Heat Shock Protein Genes During Tomato Fruit Ripening.

Heat shock proteins (HSPs) are ubiquitous and highly conserved in nature. Heat stress upregulates their gene expression and now it is known that they are also developmentally regulated. We have studied regulation of small HSP genes during ripening of tomato fruit. In this study, we identify two small HSP genes, SlHSP17.7A and SlHSP17.7B, localized on tomato Chr.6 and Chr.9, respectively. Each gene encodes proteins constituting 154 amino acids and has characteristic domains as in other sHSP genes. We found that SlHSP17.7A and SlHSP17.7B gene expression is low in the vegetative tissues as compared to that in the fruit. These sHSP genes are characteristically expressed in a fruit-ripening fashion, being upregulated during the ripening transition of mature green to breaker stage. Their expression patterns mirror that of the rate-limiting ethylene biosynthesis gene ACC (1-aminocyclopropane-1-carboxylic acid) synthase, SlACS2, and its regulator SlMADS-RIN. Exogenous application of ethylene to either mature green tomato fruit or tomato leaves suppressed the expression of both the SlHSP17.7A, B genes. Notably and characteristically, a transgenic tomato line silenced for SlACS2 gene and whose fruits produce ~50% less ethylene in vivo, had higher expression of both the sHSP genes at the fruit ripening transition stages [breaker (BR) and BR+3] than the control fruit. Moreover, differential gene expression of SlHSP17.7A versus SlHSP17.7B gene was apparent in the tomato ripening mutants-rin/rin, nor/nor, and Nr/Nr, with the expression of SlHSP17.7A being significantly reduced but that of SlHSP17.7B significantly upregulated as compared to the wild type (WT). These data indicate that ethylene negatively regulates transcriptional abundance of both these sHSPs. Transient overexpression of the ripening regulator SlMADS-RIN in WT and ACS2-AS mature green tomato fruits suppressed the expression of SlHSP17.7A but not that of SlHSP17.7B. Thus, ethylene directly or in tune with SlMADS-RIN regulates the transcript abundance of both these sHSP genes.

Transcriptional Regulation of Abscission Zones.

Precise and timely regulation of organ separation from the parent plant (abscission) is consequential to improvement of crop productivity as it influences both the timing of harvest and fruit quality. Abscission is tightly associated with plant fitness as unwanted organs (petals, sepals, filaments) are shed after fertilization while seeds, fruits, and leaves are cast off as means of reproductive success or in response to abiotic/biotic stresses. Floral organ abscission in Arabidopsis has been a useful model to elucidate the molecular mechanisms that underlie the separation processes, and multiple abscission signals associated with the activation and downstream pathways have been uncovered. Concomitantly, large-scale analyses of omics studies in diverse abscission systems of various plants have added valuable insights into the abscission process. The results suggest that there are common molecular events linked to the biosynthesis of a new extracellular matrix as well as cell wall disassembly. Comparative analysis between Arabidopsis and soybean abscission systems has revealed shared and yet disparate regulatory modules that affect the separation processes. In this review, we discuss our current understanding of the transcriptional regulation of abscission in several different plants that has improved on the previously proposed four-phased model of organ separation.

Re-evaluation of the ethylene-dependent and -independent pathways in the regulation of floral and organ abscission.

Abscission is a developmental process with important implications for agricultural practices. Ethylene has long been considered as a key regulator of the abscission process. The existence of an ethylene-independent abscission pathway, controlled by the complex of INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) peptide and the HAESA (HAE) and HAESA-like2 (HSL2) kinases, has been proposed, based mainly on observations that organ abscission in ethylene-insensitive mutants was delayed but not inhibited. A recent review on plant organ abscission signaling highlighted the IDA-HAE-HSL2 components as the regulators of organ abscission, while the role of auxin and ethylene in this process was hardly addressed. After a careful analysis of the relevant abscission literature, we propose that the IDA-HAE-HSL2 pathway is essential for the final stages of organ abscission, while ethylene plays a major role in its initiation and progression. We discuss the view that the IDA-HAE-HSL2 pathway is ethylene independent, and present recent evidence showing that ethylene activates the IDA-HAE-HSL2 complex. We conclude that the ability of an organ to abscise is tightly linked to cell turgidity in the abscission zone, and suggest that lack of cell turgidity might contribute to the failure of floral organ abscission in the ida mutants.

The root-knot nematode Meloidogyne incognita produces a functional mimic of the Arabidopsis INFLORESCENCE DEFICIENT IN ABSCISSION signaling peptide.

INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) is a signaling peptide that regulates cell separation in Arabidopsis including floral organ abscission and lateral root emergence. IDA is highly conserved in dicotyledonous flowering plant genomes. IDA-like sequences were also found in the genomic sequences of root-knot nematodes, Meloidogyne spp., which are globally deleterious pathogens of agriculturally important plants, but the role of these genes is unknown. Exogenous treatment of the Arabidopsis ida mutant with synthetic peptide identical to the M. incognita IDA-like 1 (MiIDL1) protein sequence minus its N-terminal signal peptide recovered both the abscission and root architecture defects. Constitutive expression of the full-length MiIDL1 open reading frame in the ida mutant substantially recovered the delayed floral organ abscission phenotype whereas transformants expressing a construct missing the MiIDL1 signal peptide retained the delayed abscission phenotype. Importantly, wild-type Arabidopsis plants harboring an MiIDL1-RNAi construct and infected with nematodes had approximately 40% fewer galls per root than control plants. Thus, the MiIDL1 gene produces a functional IDA mimic that appears to play a role in successful gall development on Arabidopsis roots.

Transient regulation of three clustered tomato class-I small heat-shock chaperone genes by ethylene is mediated by SlMADS-RIN transcription factor.

Clustered class-I small heat-shock protein (sHSP) chaperone genes, SlHSP17.6, SlHSP20.0 and SlHSP20.1, in tomato are demonstrated to be transcriptionally regulated by ethylene during mature green (MG) fruit transition into ripening. These genes are constitutively expressed at MG fruit stage in two different tomato genotypes as well as in their ripening mutants, including rin, nor and Nr, and an ethylene-deficient transgenic line, ACS2-antisense. Notably, ethylene treatment of the MG fruit led to significant sHSP gene suppression in both wild-types, ACS2-antisense, nor/nor and Nr/Nr, but not the rin/rin mutant. Inability of ethylene to suppress sHSP genes in rin/rin mutant, which harbors MADS-RIN gene mutation, suggests that MADS-RIN transcription factor regulates the expression of these genes. Treatment of the wild type and ACS2-antisense fruit with the ethylene-signaling inhibitor, 1-methylcyclopropane (1-MCP), reversed the sHSP gene suppression. Transcripts of representative ethylene-responsive and ripening-modulated genes confirmed and validated sHSP transcript profile patterns. In silico analysis in conjunction with chromatin immunoprecipitation demonstrated MADS-RIN protein binding to specific CArG motifs present in the promoters of these chaperone genes. The results establish MADS-RIN protein as a transcriptional regulator of these chaperone genes in an ethylene-dependent manner, and that MADS-RIN protein-regulation of sHSPs is integral to tomato fruit ripening.

Treatment of Plants with Gaseous Ethylene and Gaseous Inhibitors of Ethylene Action.

The gaseous nature of ethylene affects not only its role in plant biology but also how you treat plants with the hormone. In many ways, it simplifies the treatment problem. Other hormones have to be made up in solution and applied to some part of the plant hoping the hormone will be taken up into the plant and translocated throughout the plant at the desired concentration. Because all plant cells are connected by an intercellular gas space the ethylene concentration you treat with is relatively quickly reached throughout the plant. In some instances, like mature fruit, treatment with ethylene initiates autocatalytic synthesis of ethylene. However, in most experiments, the exogenous ethylene concentration is saturating, usually >1 µL L-1, and the synthesis of additional ethylene is inconsequential. Also facilitating ethylene research compared with other hormones is that there are inhibitors of ethylene action 1-MCP (1-methylcyclopropene) and 2,5-NBD (2,5-norbornadiene) that are also gases wherein you can achieve nearly 100% inhibition of ethylene action quickly and with few side effects. Inhibitors for other plant hormones are applied as a solution and their transport and concentration at the desired site is not always known and difficult to measure. Here, our focus is on how to treat plants and plant parts with the ethylene gas and the gaseous inhibitors of ethylene action.

Transcriptome Analysis of Soybean Leaf Abscission Identifies Transcriptional Regulators of Organ Polarity and Cell Fate.

Abscission, organ separation, is a developmental process that is modulated by endogenous and environmental factors. To better understand the molecular events underlying the progression of abscission in soybean, an agriculturally important legume, we performed RNA sequencing (RNA-seq) of RNA isolated from the leaf abscission zones (LAZ) and petioles (Non-AZ, NAZ) after treating stem/petiole explants with ethylene for 0, 12, 24, 48, and 72 h. As expected, expression of several families of cell wall modifying enzymes and many pathogenesis-related (PR) genes specifically increased in the LAZ as abscission progressed. Here, we focus on the 5,206 soybean genes we identified as encoding transcription factors (TFs). Of the 5,206 TFs, 1,088 were differentially up- or down-regulated more than eight-fold in the LAZ over time, and, within this group, 188 of the TFs were differentially regulated more than eight-fold in the LAZ relative to the NAZ. These 188 abscission-specific TFs include several TFs containing domains for homeobox, MYB, Zinc finger, bHLH, AP2, NAC, WRKY, YABBY, and auxin-related motifs. To discover the connectivity among the TFs and highlight developmental processes that support organ separation, the 188 abscission-specific TFs were then clustered based on a >four-fold up- or down-regulation in two consecutive time points (i.e., 0 and 12 h, 12 and 24 h, 24 and 48 h, or 48 and 72 h). By requiring a sustained change in expression over two consecutive time intervals and not just one or several time intervals, we could better tie changes in TFs to a particular process or phase of abscission. The greatest number of TFs clustered into the 0 and 12 h group. Transcriptional network analysis for these abscission-specific TFs indicated that most of these TFs are known as key determinants in the maintenance of organ polarity, lateral organ growth, and cell fate. The abscission-specific expression of these TFs prior to the onset of abscission and their functional properties as defined by studies in Arabidopsis indicate that these TFs are involved in defining the separation cells and initiation of separation within the AZ by balancing organ polarity, roles of plant hormones, and cell differentiation.

Examination of the Abscission-Associated Transcriptomes for Soybean, Tomato, and Arabidopsis Highlights the Conserved Biosynthesis of an Extensible Extracellular Matrix and Boundary Layer.

Abscission zone (AZ) development and the progression of abscission (detachment of plant organs) have been roughly separated into four stages: first, AZ differentiation; second, competence to respond to abscission signals; third, activation of abscission; and fourth, formation of a protective layer and post-abscission trans-differentiation. Stage three, activation of abscission, is when changes in the cell wall and extracellular matrix occur to support successful organ separation. Most abscission research has focused on gene expression for enzymes that disassemble the cell wall within the AZ and changes in phytohormones and other signaling events that regulate their expression. Here, transcriptome data for soybean, tomato and Arabidopsis were examined and compared with a focus not only on genes associated with disassembly of the cell wall but also on gene expression linked to the biosynthesis of a new extracellular matrix. AZ-specific up-regulation of genes associated with cell wall disassembly including cellulases (beta-1,4-endoglucanases, CELs), polygalacturonases (PGs), and expansins (EXPs) were much as expected; however, curiously, changes in expression of xyloglucan endotransglucosylase/hydrolases (XTHs) were not AZ-specific in soybean. Unexpectedly, we identified an early increase in the expression of genes underlying the synthesis of a waxy-like cuticle. Based on the expression data, we propose that the early up-regulation of an abundance of small pathogenesis-related (PR) genes is more closely linked to structural changes in the extracellular matrix of separating cells than an enzymatic role in pathogen resistance. Furthermore, these observations led us to propose that, in addition to cell wall loosening enzymes, abscission requires (or is enhanced by) biosynthesis and secretion of small proteins (15-25 kDa) and waxes that form an extensible extracellular matrix and boundary layer on the surface of separating cells. The synthesis of the boundary layer precedes what is typically associated with the post-abscission synthesis of a protective scar over the fracture plane. This modification in the abscission model is discussed in regard to how it influences our interpretation of the role of multiple abscission signals.

To grow old: regulatory role of ethylene and jasmonic acid in senescence.

Senescence, the final stage in the development of an organ or whole plant, is a genetically programmed process controlled by developmental and environmental signals. Age-related signals underlie the onset of senescence in specific organs (leaf, flower, and fruit) as well as the whole plant (monocarpic senescence). Rudimentary to most senescence processes is the plant hormone ethylene, a small gaseous molecule critical to diverse processes throughout the life of the plant. The role of ethylene in senescence was discovered almost 100 years ago, but the molecular mechanisms by which ethylene regulates senescence have been deciphered more recently primarily through genetic and molecular studies in Arabidopsis. Jasmonic acid (JA), another plant hormone, is emerging as a key player in the control of senescence. The regulatory network of ethylene and JA involves the integration of transcription factors, microRNAs, and other hormones. In this review, we summarize the current understanding of ethylene's role in senescence, and discuss the interplay of ethylene with JA in the regulation of senescence.

Abscission of flowers and floral organs is closely associated with alkalization of the cytosol in abscission zone cells.

In vivo changes in the cytosolic pH of abscission zone (AZ) cells were visualized using confocal microscopic detection of the fluorescent pH-sensitive and intracellularly trapped dye, 2',7'-bis-(2-carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF), driven by its acetoxymethyl ester. A specific and gradual increase in the cytosolic pH of AZ cells was observed during natural abscission of flower organs in Arabidopsis thaliana and wild rocket (Diplotaxis tenuifolia), and during flower pedicel abscission induced by flower removal in tomato (Solanum lycopersicum Mill). The alkalization pattern in the first two species paralleled the acceleration or inhibition of flower organ abscission induced by ethylene or its inhibitor 1-methylcyclopropene (1-MCP), respectively. Similarly, 1-MCP pre-treatment of tomato inflorescence explants abolished the pH increase in AZ cells and pedicel abscission induced by flower removal. Examination of the pH changes in the AZ cells of Arabidopsis mutants defective in both ethylene-induced (ctr1, ein2, eto4) and ethylene-independent (ida, nev7, dab5) abscission pathways confirmed these results. The data indicate that the pH changes in the AZ cells are part of both the ethylene-sensitive and -insensitive abscission pathways, and occur concomitantly with the execution of organ abscission. pH can affect enzymatic activities and/or act as a signal for gene expression. Changes in pH during abscission could occur via regulation of transporters in AZ cells, which might affect cytosolic pH. Indeed, four genes associated with pH regulation, vacuolar H(+)-ATPase, putative high-affinity nitrate transporter, and two GTP-binding proteins, were specifically up-regulated in tomato flower AZ following abscission induction, and 1-MCP reduced or abolished the increased expression.

De novo Transcriptome Sequencing and Development of Abscission Zone-Specific Microarray as a New Molecular Tool for Analysis of Tomato Organ Abscission.

Abscission of flower pedicels and leaf petioles of tomato (Solanum lycopersicum) can be induced by flower removal or leaf deblading, respectively, which leads to auxin depletion, resulting in increased sensitivity of the abscission zone (AZ) to ethylene. However, the molecular mechanisms that drive the acquisition of abscission competence and its modulation by auxin gradients are not yet known. We used RNA-Sequencing (RNA-Seq) to obtain a comprehensive transcriptome of tomato flower AZ (FAZ) and leaf AZ (LAZ) during abscission. RNA-Seq was performed on a pool of total RNA extracted from tomato FAZ and LAZ, at different abscission stages, followed by de novo assembly. The assembled clusters contained transcripts that are already known in the Solanaceae (SOL) genomics and NCBI databases, and over 8823 identified novel tomato transcripts of varying sizes. An AZ-specific microarray, encompassing the novel transcripts identified in this study and all known transcripts from the SOL genomics and NCBI databases, was constructed to study the abscission process. Multiple probes for longer genes and key AZ-specific genes, including antisense probes for all transcripts, make this array a unique tool for studying abscission with a comprehensive set of transcripts, and for mining for naturally occurring antisense transcripts. We focused on comparing the global transcriptomes generated from the FAZ and the LAZ to establish the divergences and similarities in their transcriptional networks, and particularly to characterize the processes and transcriptional regulators enriched in gene clusters that are differentially regulated in these two AZs. This study is the first attempt to analyze the global gene expression in different AZs in tomato by combining the RNA-Seq technique with oligonucleotide microarrays. Our AZ-specific microarray chip provides a cost-effective approach for expression profiling and robust analysis of multiple samples in a rapid succession.

The haustorial transcriptomes of Uromyces appendiculatus and Phakopsora pachyrhizi and their candidate effector families.

Haustoria of biotrophic rust fungi are responsible for the uptake of nutrients from their hosts and for the production of secreted proteins, known as effectors, which modulate the host immune system. The identification of the transcriptome of haustoria and an understanding of the functions of expressed genes therefore hold essential keys for the elucidation of fungus-plant interactions and the development of novel fungal control strategies. Here, we purified haustoria from infected leaves and used 454 sequencing to examine the haustorial transcriptomes of Phakopsora pachyrhizi and Uromyces appendiculatus, the causal agents of soybean rust and common bean rust, respectively. These pathogens cause extensive yield losses in their respective legume crop hosts. A series of analyses were used to annotate expressed sequences, including transposable elements and viruses, to predict secreted proteins from the assembled sequences and to identify families of candidate effectors. This work provides a foundation for the comparative analysis of haustorial gene expression with further insights into physiology and effector evolution.

A gene encoding a peptide with similarity to the plant IDA signaling peptide (AtIDA) is expressed most abundantly in the root-knot nematode (Meloidogyne incognita) soon after root infection.

Small peptides play important roles in intercellular signaling. Inflorescence deficient in abscission (ida) is an Arabidopsis mutant that does not abscise (shed) its flower petals. The IDA gene encodes a small, secreted peptide that putatively binds to two redundant receptor-like kinases (HAESA and HAESA-like2) that initiate a signal transduction pathway. We identified IDA-like (IDL) genes in the genomic sequence for Meloidogyne incognita and Meloidogyne hapla. No orthologous sequences were found in any other genus of nematodes. Transcript for both M. incognita and M. hapla IDLs were found in total RNA isolated from infected root systems of tomato, Solanum lycopersicum. Five and three prime RACE of RNA from M. incognita infected tomato roots revealed a sequence of 392 nt that includes a poly (A) tail of 39 nt. The open reading frame encodes a 47 aa protein with a putative 25 aa N-terminal signal peptide. Expression of MiIDL1 is very low in eggs and pre-parasitic J2 and rapidly increases in the first four days post inoculation (dpi) and then declines at approximately 14 dpi. A proposed role for the root-knot nematode IDL is discussed.

CTR1 phosphorylates the central regulator EIN2 to control ethylene hormone signaling from the ER membrane to the nucleus in Arabidopsis.

The gaseous phytohormone ethylene C(2)H(4) mediates numerous aspects of growth and development. Genetic analysis has identified a number of critical elements in ethylene signaling, but how these elements interact biochemically to transduce the signal from the ethylene receptor complex at the endoplasmic reticulum (ER) membrane to transcription factors in the nucleus is unknown. To close this gap in our understanding of the ethylene signaling pathway, the challenge has been to identify the target of the CONSTITUTIVE TRIPLE RESPONSE1 (CTR1) Raf-like protein kinase, as well as the molecular events surrounding ETHYLENE-INSENSITIVE2 (EIN2), an ER membrane-localized Nramp homolog that positively regulates ethylene responses. Here we demonstrate that CTR1 interacts with and directly phosphorylates the cytosolic C-terminal domain of EIN2. Mutations that block the EIN2 phosphorylation sites result in constitutive nuclear localization of the EIN2 C terminus, concomitant with constitutive activation of ethylene responses in Arabidopsis. Our results suggest that phosphorylation of EIN2 by CTR1 prevents EIN2 from signaling in the absence of ethylene, whereas inhibition of CTR1 upon ethylene perception is a signal for cleavage and nuclear localization of the EIN2 C terminus, allowing the ethylene signal to reach the downstream transcription factors. These findings significantly advance our understanding of the mechanisms underlying ethylene signal transduction.

Proteomic pleiotropy of OpgGH, an operon necessary for efficient growth of Salmonella enterica serovar typhimurium under low-osmotic conditions.

Salmonella enterica, a bacterial, food-borne pathogen of humans, can contaminate raw fruits and vegetables. Unfortunately for consumers, the bacteria can survive in water used to wash away contaminating bacteria. The ability to survive the low-osmotic conditions of the wash water is attributed to the OpgGH operon that leads to the production of osmotically regulated periplasmic glucans. Mutants lacking OpgGH grow slowly under low-osmotic conditions, but there are also unexpected traits such as abnormal flagellar motility and reduced virulence in mice. To get a broader understanding of these pleiotropic effects under low osmolarity, we examined the proteome of these mutants using high-throughput mass spectrometry. We identified approximately one-third of the proteins encoded by the genome and used label-free spectral counting to determine the relative amounts of proteins in wild-type cultures and mutants. Mutants had reduced amounts of proteins required for osmotic sensing, flagellar motility, purine and pyrimidine metabolism, oxidative energy production, and protein translation. By contrast, mutants had greater amounts of ABC transporters needed to balance cellular osmolarity. Hence, the effects of OpgGH reach across the proteome, and the data are consistent with the mutant phenotypes.

IDA-like gene expression in soybean and tomato leaf abscission and requirement for a diffusible stelar abscission signal.

BACKGROUND AND AIMS: The stimulatory and inhibitory role of ethylene and auxin, respectively, in leaf abscission (leaf drop) is well documented. More recently, IDA (INFLORESCENCE DEFICIENT IN ABSCISSION) peptides and their putative interacting receptor-like-kinase partners, HAESA and HAESA-like2, were shown to be essential components in Arabidopsis floral organ abscission. Prior to research on IDA, it was reported that bean (Phaseolus vulgaris) leaf abscission required a diffusible signal that emanated from the vascular tissue. We were interested in determining whether the IDA signalling path might regulate abscission in plants other than Arabidopsis and whether IDA might act as a diffusible signal in abscission. METHODOLOGY: Quantitative polymerase chain reaction was used to monitor gene expression and a GUS reporter gene construct used to determine the need for a diffusible signal in tomato. PRINCIPAL RESULTS: We identified 12 IDA-like and 11 HAESA-like genes in soybean (Glycine max) and monitored their gene expression in abscission in relation to the expression of several cell-wall-modifying proteins and aminocyclopropane-1-carboxylic acid synthases. Ethylene evoked the expression of several IDA-like genes in abscission zones (AZ), but also to a lesser degree in the adjacent petiole tissue. Surprisingly, IDA-like gene expression was very high in senescent soybean leaves. We identified five IDA-like genes in tomato (Solanum lycopersicum). Only one IDA-like gene was expressed in the tomato AZ and its expression was approximately equal in the AZ and petioles, but no IDA-like gene showed significant expression in leaves at up to 96 h of exposure to ethylene. CONCLUSIONS: IDA-like gene expression is up-regulated during soybean and tomato abscission but up-regulation was not limited to the AZ. Cell separation in the AZ cortex of tomato does not require a diffusible signal emanating from the stele. A role for IDA in soybean and tomato leaf abscission is discussed.

Proteomic responses in Arabidopsis thaliana seedlings treated with ethylene.

Ethylene (ET) is a volatile hormone that modulates fruit ripening, plant growth, development and stress responses. Key components of the ET-signaling pathway identified by genetic dissection in Arabidopsis thaliana include five ET receptors, the negative regulator CTR1 and the positive regulator EIN2, all of which localize to the endoplasmic reticulum. Mechanisms of signaling among these proteins are still unresolved and targets of ET responses are not fully known. So, we used mass spectrometry to identify proteins in microsomal membrane preparations from etiolated A. thaliana seedlings maintained in ambient air or treated with ET for 3 h. We compared 3814 proteins from ET-exposed seedlings and controls and identified 304 proteins with significant accumulation changes. The proteins with increased accumulation were involved in ET biosynthesis, cell morphogenesis, oxidative stress and vesicle secretion while those with decreased accumulation were ribosomal proteins and proteins positively regulated by brassinosteroid, another hormone involved in cell elongation. Several proteins, including EIN2, appeared to be differentially phosphorylated upon ET treatment, which suggests that the activity or stability of these proteins may be controlled by phosphorylation. TUA3, a component of microtubules that contributes to cellular morphological change, exhibited both increased accumulation and differential phosphorylation upon ET treatment. To verify the role of TUA3 in the ET response, tua3 mutants were evaluated. Mutant seedlings had altered ET-associated growth movements. The data indicate that ET perception leads to rapid proteomic change and that these changes are an important part of signaling and development. The data serve as a foundation for exploring ET signaling through systems biology.

Gene expression profiling and shared promoter motif for cell wall-modifying proteins expressed in soybean cyst nematode-infected roots.

We hypothesized that soybean cyst nematode (SCN; Heterodera glycines) co-opts part or all of one or more innate developmental process in soybean (Glycine max) to establish its feeding structure, syncytium, in soybean roots. The syncytium is formed within the vascular bundle by partial degradation of cell walls and membranes between adjacent parenchyma cells. A mature syncytium incorporates as many as 200 cells into one large multinucleated cell. Gene expression patterns for several cell wall-modifying proteins were compared in multiple tissues undergoing major shifts in cell wall integrity. These included SCN-colonized roots, root tips where vascular differentiation occurs, flooded roots (aerenchyma), adventitious rooting in hypocotyls, and leaf abscission zones. A search in the 5' upstream promoters of these genes identified a motif (SCNbox1: WGCATGTG) common to several genes that were up-regulated in several different tissues. The polygalacturonase 11 promoters (GmPG11a/b) include the SCNbox1 motif. The expression pattern for GmPG11a was examined further in transgenic soybean containing a PG11a promoter fused to a ß-glucuronidase (GUS) reporter gene. GUS expression was highest in cells undergoing radial expansion in the stele and/or cell wall dissolution. GUS staining was not observed in cortical cells where a lateral root tip or a growing nematode emerged through the root cortex.