Plant Gene Expression Center

The basic/helix-loop-helix (bHLH) proteins are a superfamily of transcription factors that bind as dimers to specific DNA target sites and that have been well characterized in nonplant eukaryotes as important regulatory components in diverse biological processes. Based on evidence that the bHLH protein PIF3 is a direct phytochrome reaction partner in the photoreceptor's signaling network, we have undertaken a comprehensive computational analysis of the Arabidopsis genome sequence databases to define the scope and features of the bHLH family. Using a set of criteria derived from a previously defined consensus motif, we identified 147 bHLH protein-encoding genes, making this one of the largest transcription factor families in Arabidopsis. Phylogenetic analysis of the bHLH domain sequences permits classification of these genes into 21 subfamilies. The evolutionary and potential functional relationships implied by this analysis are supported by other criteria, including the chromosomal distribution of these genes relative to duplicated genome segments, the conservation of variant exon/intron structural patterns, and the predicted DNA binding activities within subfamilies. Considerable diversity in DNA binding site specificity among family members is predicted, and marked divergence in protein sequence outside of the conserved bHLH domain is observed. Together with the established propensity of bHLH factors to engage in varying degrees of homodimerization and heterodimerization, these observations suggest that the Arabidopsis bHLH proteins have the potential to participate in an extensive set of combinatorial interactions, endowing them with the capacity to be involved in the regulation of a multiplicity of transcriptional programs. We provide evidence from yeast two-hybrid and in vitro binding assays that two related phytochrome-interacting members in the Arabidopsis family, PIF3 and PIF4, can form both homodimers and heterodimers and that all three dimeric configurations can bind specifically to the G-box DNA sequence motif CACGTG. These data are consistent, in principle, with the operation of this combinatorial mechanism in Arabidopsis.

TRAIL is a tumor necrosis factor-related ligand that induces apoptosis upon binding to its death domain-containing receptors, DR4 and DR5. Two additional TRAIL receptors, TRID/DcR1 and DcR2, lack functional death domains and function as decoy receptors for TRAIL. We have identified a fifth TRAIL receptor, namely osteoprotegerin (OPG), a secreted tumor necrosis factor receptor homologue that inhibits osteoclastogenesis and increases bone density in vivo. OPG-Fc binds TRAIL with an affinity of 3.0 nM, which is slightly weaker than the interaction of TRID-Fc or DR5-Fc with TRAIL. OPG inhibits TRAIL-induced apoptosis of Jurkat cells. Conversely, TRAIL blocks the anti-osteoclastogenic activity of OPG. These data suggest potential cross-regulatory mechanisms by OPG and TRAIL.

The AUXIN RESPONSE FACTOR (ARF) gene family products, together with the AUXIN/INDOLE-3-ACETIC ACID proteins, regulate auxin-mediated transcriptional activation/repression. The biological function(s) of most ARFs is poorly understood. Here, we report the identification and characterization of T-DNA insertion lines for 18 of the 23 ARF gene family members in Arabidopsis thaliana. Most of the lines fail to show an obvious growth phenotype except of the previously identified arf2/hss, arf3/ett, arf5/mp, and arf7/nph4 mutants, suggesting that there are functional redundancies among the ARF proteins. Subsequently, we generated double mutants. arf7 arf19 has a strong auxin-related phenotype not observed in the arf7 and arf19 single mutants, including severely impaired lateral root formation and abnormal gravitropism in both hypocotyl and root. Global gene expression analysis revealed that auxin-induced gene expression is severely impaired in the arf7 single and arf7 arf19 double mutants. For example, the expression of several genes, such as those encoding members of LATERAL ORGAN BOUNDARIES domain proteins and AUXIN-REGULATED GENE INVOLVED IN ORGAN SIZE, are disrupted in the double mutant. The data suggest that the ARF7 and ARF19 proteins play essential roles in auxin-mediated plant development by regulating both unique and partially overlapping sets of target genes. These observations provide molecular insight into the unique and overlapping functions of ARF gene family members in Arabidopsis.

Abstract Lateral root formation in Arabidopsis thaliana is regulated by two related AUXIN RESPONSE FACTORs, ARF7 and ARF19, which are transcriptional activators of early auxin response genes. The arf7 arf19 double knockout mutant is severely impaired in lateral root formation. Target-gene analysis in arf7 arf19 transgenic plants harboring inducible forms of ARF7 and ARF19 revealed that ARF7 and ARF19 directly regulate the auxin-mediated transcription of LATERAL ORGAN BOUNDARIES-DOMAIN16/ASYMMETRIC LEAVES2-LIKE18 (LBD16/ASL18) and/or LBD29/ASL16 in roots. Overexpression of LBD16/ASL18 and LBD29/ASL16 induces lateral root formation in the absence of ARF7 and ARF19. These LBD/ASL proteins are localized in the nucleus, and dominant repression of LBD16/ASL18 activity inhibits lateral root formation and auxin-mediated gene expression, strongly suggesting that these LBD/ASLs function downstream of ARF7- and ARF19-dependent auxin signaling in lateral root formation. Our results reveal that ARFs regulate lateral root formation via direct activation of LBD/ASLs in Arabidopsis.

We often think of plants primarily as a source of wood, food, and fiber. Secondarily we may also appreciate their presence for aesthetic reasons as well as for altruistically providing habitat for other species. Increasingly, however, their value as an environmental counterbalance to industrialization processes is being appreciated. These processes include the burning of fossil fuels, generation of wastes (sewage, inorganic and organic solids, and effluents), and general water flow and processing. Plants have long been recognized for their consumption of CO2 and, more recently, of other gaseous industrial byproducts (Simonich and Hites, 1994). Recently, their role in slowing the rate of global warming has been further appreciated in both the scientific and popular press. Their use as a final water treatment and for disposal of sludge resulting from waste water treatment is centuries old (Hartman, 1975). The extensive literature concerning water and sludge treatment and the emerging field of air pollution abatement with plants will not be discussed here. Instead, we focus on an emerging concept, phytoremediation, the use of plants to remediate contamination of soil with organic or inorganic wastes. Remediation of soil contamination by conventional engineering techniques often costs between $50 and $500 per ton. Certain specialized techniques can exceed costs of $1000 per ton. With an acre of soil (to a 3-foot depth) weighing approximately 4500 tons, this translates to a minimum cost of about a quarter million dollars per acre (Cunningham et al., 1995). It is not surprising that the cleanup of contaminated sites has not been proceeding at a rapid pace. There is an active effort to develop new, more costeffective technologies to remediate contamination of such soils. For the most part these efforts are being led by engineers and microbiologists. More recently, however, green plant-based processes have begun receiving greater attention. It has long been known that the life cycle of a plant has profound effects on the chemical, physical, and biological processes that occur in its immediate vicinity. In the process of shoot and root growth, water and mineral acquisition, senescence, and eventual decay, plants can profoundly alter the surrounding soil. The effects of many of these processes are apparent on the restoration of land at physically and chemically altered sites, ranging from road cuts to the site of the Mount St. Helen's eruption. These same plant-driven processes also occur in areas heavily impacted by industrial, mining, and urban activities. One of the greatest forces driving increased emphasis on research in this area is the potential economic benefit of an agronomybased technology. Growing a crop on an acre of land can be accomplished at a cost ranging from 2 to 4 orders of magnitude less than the current engineering cost of excavation and reburial. There have been perhaps two dozen field tests to date; however, in many ways phytoremediation is still at its initial stages of research and development. A comforting thought for plant biologists is that much of the research effort will be expected to center on a deeper understanding of basic plant processes. So how do we envision phytoremediation working? The theory appears to be simple. Agronomic techniques will be used to ready the contaminated soil for planting and to ameliorate chemical and physical limitations to plant growth. Plants will then directly or indirectly absorb, sequester, and/or degrade the contaminant. Plants and irrigation, fertilization, and cropping schemes will be managed to maximize this remedial effect. By growing plants over a number of years, the aim is to either remove the pollutant from the contaminated matrix or to alter the chemical and physical nature of the contaminant within the soil so that it no longer presents a risk to human health and the environment. As people who work in the remediation, herbicide development, and farming industries will attest, many weed species are remarkably tolerant of a wide range of organic and inorganic toxins. Plants can thrive in soil contaminated to levels that are often orders of magnitude higher than current regulatory limits. These limits are often set relatively independent of plant tolerance limits and are most often derived from human health and aquatic toxicology end points. Ironically, many remediation plans begin with the destruction of the existing vegetation.

Genome wide analysis of orthologous clusters is an important component of comparative genomics studies. Identifying the overlap among orthologous clusters can enable us to elucidate the function and evolution of proteins across multiple species. Here, we report a web platform named OrthoVenn that is useful for genome wide comparisons and visualization of orthologous clusters. OrthoVenn provides coverage of vertebrates, metazoa, protists, fungi, plants and bacteria for the comparison of orthologous clusters and also supports uploading of customized protein sequences from user-defined species. An interactive Venn diagram, summary counts, and functional summaries of the disjunction and intersection of clusters shared between species are displayed as part of the OrthoVenn result. OrthoVenn also includes in-depth views of the clusters using various sequence analysis tools. Furthermore, OrthoVenn identifies orthologous clusters of single copy genes and allows for a customized search of clusters of specific genes through key words or BLAST. OrthoVenn is an efficient and user-friendly web server freely accessible at http://probes.pw.usda.gov/OrthoVenn or http://aegilops.wheat.ucdavis.edu/OrthoVenn.

Plant breeders have used disease resistance genes (R genes) to control plant disease since the turn of the century. Molecular cloning of R genes that enable plants to resist a diverse range of pathogens has revealed that the proteins encoded by these genes have several features in common. These findings suggest that plants may have evolved common signal transduction mechanisms for the expression of resistance to a wide range of unrelated pathogens. Characterization of the molecular signals involved in pathogen recognition and of the molecular events that specify the expression of resistance may lead to novel strategies for plant disease control.

Abstract OrthoVenn is a powerful web platform for the comparison and analysis of whole-genome orthologous clusters. Here we present an updated version, OrthoVenn2, which provides new features that facilitate the comparative analysis of orthologous clusters among up to 12 species. Additionally, this update offers improvements to data visualization and interpretation, including an occurrence pattern table for interrogating the overlap of each orthologous group for the queried species. Within the occurrence table, the functional annotations and summaries of the disjunctions and intersections of clusters between the chosen species can be displayed through an interactive Venn diagram. To facilitate a broader range of comparisons, a larger number of species, including vertebrates, metazoa, protists, fungi, plants and bacteria, have been added in OrthoVenn2. Finally, a stand-alone version is available to perform large dataset comparisons and to visualize results locally without limitation of species number. In summary, OrthoVenn2 is an efficient and user-friendly web server freely accessible at https://orthovenn2.bioinfotoolkits.net.

Functional analysis of a genome requires accurate gene structure information and a complete gene inventory. A dual experimental strategy was used to verify and correct the initial genome sequence annotation of the reference plant Arabidopsis. Sequencing full-length cDNAs and hybridizations using RNA populations from various tissues to a set of high-density oligonucleotide arrays spanning the entire genome allowed the accurate annotation of thousands of gene structures. We identified 5817 novel transcription units, including a substantial amount of antisense gene transcription, and 40 genes within the genetically defined centromeres. This approach resulted in completion of approximately 30% of the Arabidopsis ORFeome as a resource for global functional experimentation of the plant proteome.

Light signals are fundamental to the growth and development of plants. Red and far-red light are sensed using the phytochrome family of plant photoreceptors. Individual phytochromes display both unique and overlapping roles throughout the life cycle of plants, regulating a range of developmental processes from seed germination to the timing of reproductive development. The evolution of multiple phytochrome photoreceptors has enhanced plant sensitivity to fluctuating light environments, diversifying phytochrome function, and facilitating conditional cross-talk with other signalling systems. The isolation of null mutants, deficient in all individual phytochromes, has greatly advanced understanding of phytochrome functions in the model species, Arabidopsis thaliana. The creation of mutants null for multiple phytochrome combinations has enabled the dissection of redundant interactions between family members, revealing novel regulatory roles for this important photoreceptor family. In this review, current knowledge of phytochrome functions in the light-regulated development of Arabidopsis is summarised.

Phytochrome is a plant regulatory photoreceptor that mediates red light effects on a wide variety of physiological and molecular responses. DNA blot analysis indicates that the Arabidopsis thaliana genome contains four to five phytochrome-related gene sequences. We have isolated and sequenced cDNA clones corresponding to three of these genes and have deduced the amino acid sequence of the full-length polypeptide encoded in each case. One of these proteins (phyA) shows 65-80% amino acid sequence identity with the major, etiolated-tissue phytochrome apoproteins described previously in other plant species. The other two polypeptides (phyB and phyC) are unique in that they have low sequence identity (approximately 50%) with each other, with phyA, and with all previously described phytochromes. The phyA, phyB, and phyC proteins are of similar molecular mass, have related hydropathic profiles, and contain a conserved chromophore attachment region. However, the sequence comparison data indicate that the three phy genes diverged early in plant evolution, well before the divergence of the two major groups of angiosperms, the monocots and dicots. The steady-state level of the phyA transcript is high in dark-grown A. thaliana seedlings and is down-regulated by light. In contrast, the phyB and phyC transcripts are present at lower levels and are not strongly light-regulated. These findings indicate that the red/far light-responsive phytochrome photoreceptor system in A. thaliana, and perhaps in all higher plants, consists of a family of chromoproteins that are heterogeneous in structure and regulation.

Analysis of viral and bacterial pathogenesis has revealed common themes in the ways in which plants and animals respond to pathogenic agents. Pathogenic bacteria use macromolecule delivery systems (types III and IV) to deliver microbial avirulence proteins and transfer DNA-protein complexes directly into plant cells. The molecular events that constitute critical steps of plant-pathogen interactions seem to involve ligand-receptor mechanisms for pathogen recognition and the induction of signal transduction pathways in the plant that lead to defense responses. Unraveling the molecular basis of disease resistance pathways has laid a foundation for the rational design of crop protection strategies.

Ethylene controls fruit ripening. Expression of antisense RNA to the rate-limiting enzyme in the biosynthetic pathway of ethylene, 1-aminocyclopropane-1-carboxylate synthase, inhibits fruit ripening in tomato plants. Administration of exogenous ethylene or propylene reverses the inhibitory effect. This result demonstrates that ethylene is the trigger and not the by-product of ripening and raises the prospect that the life-span of plant tissues can be extended, thereby preventing spoilage.

ABSTRACT In this paper we describe the expression patterns of a family of homeobox genes in maize and their relationship to organogenic domains in the vegetative shoot apical meristem. These genes are related by sequence to KNOTTED1, a gene characterized by dominant neomorphic mutations which perturb specific aspects of maize leaf development. Four members of this gene family are expressed in shoot meristems and the developing stem, but not in determinate lateral organs such as leaves or floral organs. The genes show distinct expression patterns in the vegetative shoot apical meristem that together predict the site of leaf initiation and the basal limit of the vegetative ‘phytomer’ or segmentation unit of the shoot. These genes are also expressed in the inflorescence and floral meristems, where their patterns of expression are more similar, and they are not expressed in root apical meristems. These findings are discussed in relation to other studies of shoot apical meristem organization as well as possible commonality of homeobox gene function in the animal and plant kingdoms.

The plant hormone IAA (or auxin) is central to the control of plant growth and development. Processes governed by auxin in concert with other plant growth regulators include development of vascular tissues, formation of lateral and adventitious roots, control of apical dominance, and tropic responses (Went and Thimann, 1937). At the level of cellular physiology, auxin profoundly affects turgor, elongation, division, and cell differentiation, the major driving and shaping forces in morphogenesis and oncogenesis. The molecular mechanisms of auxin action are still unknown, although it is now well established that auxin modulates membrane function and gene expression (for review, see Napier and Venis, 1995). These biochemical changes, in turn, most likely affect fundamental aspects of plant morphology and physiology. However, a causal relationship between auxin-mediated alterations in gene expression or membrane function and a particular growth process has not yet been demonstrated. Despite its critical role in plant development and the immense volume of studies on the diverse auxin effects, understanding of the molecular mechanisms of auxin action remains one of the major challenges in plant biology. The signal transduction cascades leading from auxin perception to altered gene expression or membrane function hold the key in our attempts to elucidate the primary mechanism(s) of auxin action. An array of experimental strategies has been mounted to investigate auxin signaling pathways. The combination of biochemical, molecular, and genetic approaches will allow for significant new insights into how the hormone works in molecular terms (Fig. 1). One strategy employs genetics and reverse genetics to construct transgenic plants with perturbations in auxin homeostasis and to screen for mutants with defects in auxinrelated physiology. Transgenic plants expressing altered hormone levels have already resolved some longstanding questions in plant physiology. Mutant plants defective in auxin responses will rejuvenate and stimulate research by identifying novel genes involved in hormone perception, signal transduction, and physiological responses (for review, see Hobbie and Estelle, 1994; Klee and Romano, 1994). The first significant result (to our knowledge) of this approach was the cloning of the AXR1 gene, which encodes a protein related to the ubiquitin-activating enzyme El (Leyser et al., 1993). Although AXRl is probably not a functional El homolog, it is nonetheless an exquisite example of the potential of molecular genetics to connect the unexpected. The biochemical strategy is based on the identification of auxin receptors and subsequent isolation of interacting components. The search for auxin receptors has led to the discovery of a number of soluble and membranebound proteins that bind auxin with moderate but physiologically relevant affinity. Their functional role in auxin signaling is still unclear and is a major target of current research (for review, see Jones, 1994; Napier and Venis, 1995). Auxin-regulated genes provide yet another source of molecular tools to dissect auxin action. The hormone modulates gene expression in a wide variety of plant tissues and cell types over a broad period of time (for review, see Guilfoyle, 1986; Theologis, 1986). However, early genes selectively induced as a primary response to auxin and prior to the initiation of cell growth are likely candidates to play a pivotal role in mediating growth-stimulating effects of the hormone. This review focuses on recent advances in our knowledge on early auxin-inducible gene expression and possible functions of the polypeptides encoded.

The phytochrome family of photoreceptors monitors the light environment and dictates patterns of gene expression that enable the plant to optimize growth and development in accordance with prevailing conditions. The enduring challenge is to define the biochemical mechanism of phytochrome action and to dissect the signaling circuitry by which the photoreceptor molecules relay sensory information to the genes they regulate. Evidence indicates that individual phytochromes have specialized photosensory functions. The amino-terminal domain of the molecule determines this photosensory specificity, whereas a short segment in the carboxyl-terminal domain is critical for signal transfer to downstream components. Heterotrimeric GTP-binding proteins, calcium-calmodulin, cyclic guanosine 5'-phosphate, and the COP-DET-FUS class of master regulators are implicated as signaling intermediates in phototransduction.

Root-associated bacterial communities play a vital role in maintaining health of the plant host. These communities exist in complex relationships, where composition and abundance of community members is dependent on a number of factors such as local soil chemistry, plant genotype and phenotype, and perturbations in the surrounding abiotic environment. One common perturbation, drought, has been shown to have drastic effects on bacterial communities, yet little is understood about the underlying causes behind observed shifts in microbial abundance. As drought may affect root bacterial communities both directly by modulating moisture availability, as well as indirectly by altering soil chemistry and plant phenotypes, we provide a synthesis of observed trends in recent studies and discuss possible directions for future research that we hope will provide for more knowledgeable predictions about community responses to future drought events.

Desert plants are hypothesized to survive the environmental stress inherent to these regions in part thanks to symbioses with microorganisms, and yet these microbial species, the communities they form, and the forces that influence them are poorly understood. Here we report the first comprehensive investigation of the microbial communities associated with species of Agave, which are native to semiarid and arid regions of Central and North America and are emerging as biofuel feedstocks. We examined prokaryotic and fungal communities in the rhizosphere, phyllosphere, leaf and root endosphere, as well as proximal and distal soil samples from cultivated and native agaves, through Illumina amplicon sequencing. Phylogenetic profiling revealed that the composition of prokaryotic communities was primarily determined by the plant compartment, whereas the composition of fungal communities was mainly influenced by the biogeography of the host species. Cultivated A. tequilana exhibited lower levels of prokaryotic diversity compared with native agaves, although no differences in microbial diversity were found in the endosphere. Agaves shared core prokaryotic and fungal taxa known to promote plant growth and confer tolerance to abiotic stress, which suggests common principles underpinning Agave-microbe interactions.

Plasmodesmata are intercellular organelles in plants that establish cytoplasmic continuity between neighboring cells. Microinjection studies showed that plasmodesmata facilitate the cell-to-cell transport of a plant-encoded transcription factor, KNOTTED1 (KN1). KN1 can also mediate the selective plasmodesmal trafficking of kn1 sense RNA. The emerging picture of plant development suggests that cell fate is determined at least in part by supracellular controls responding to cellular position as well as lineage. One of the mechanisms that enables the necessary intercellular communication appears to involve transfer of informational molecules (proteins and RNA) through plasmodesmata.

Light signals perceived by the phytochrome family of sensory photoreceptors are transduced to photoresponsive genes by an unknown mechanism. Here, we show that the basic helix-loop-helix transcription factor PIF3 binds specifically to a G-box DNA-sequence motif present in various light-regulated gene promoters, and that phytochrome B binds reversibly to G-box-bound PIF3 specifically upon light-triggered conversion of the photoreceptor to its biologically active conformer. We suggest that the phytochromes may function as integral light-switchable components of transcriptional regulator complexes, permitting continuous and immediate sensing of changes in this environmental signal directly at target gene promoters.