Characterization of Adaptation in Phototropism of Arabidopsis thaliana

Phototropic curvature has been measured for etiolated Arabidopsis thaliana seedlings with and without a preirradiation. A bilateral preirradiation with 450-nm light at a fluence greater than about 0.1 micromole per square meter causes a rapid desensitization to a subsequent 450-nanometer unilateral irradiation at 0.5 micromole per square meter. Following a refractory period, the capacity to respond phototropically recovers to the predesensitization level, and the response is then enhanced. The length of the refractory period is between 10 and 20 minutes. Both the time needed for recovery and the extent of enhancement increase with increasing fluence of the bilateral preirradiation. Based on the relative spectral sensitivities of desensitization and enhancement, these responses can be separated. Desensitization is induced by blue light but not by red light. Enhancement, however, is induced by both blue and red light. Thus, enhancement can be induced without desensitization but not vice versa. Both desensitization and enhancement affect only the magnitude of the response and do not affect the fluence threshold.     

Growth Distribution during Phototropism of Arabidopsis thaliana Seedlings

The elongation rates of two opposite sides of hypocotyls of Arabidopsis thaliana seedlings were measured during phototropism by using an infrared imaging system. In first positive phototropism, second positive phototropism, and red light-enhanced first positive phototropism, curvature toward the light source was the result of an increase in the rate of elongation of the shaded side and a decrease in the rate of elongation of the lighted side of the seedlings. The phase of straightening that followed maximum curvature resulted from a decrease in the elongation rate of the shaded side and an increase in the elongation rate of the lighted side. These data for the three types of blue light-induced phototropism tested in this study and for the phase of straightening are all clearly consistent with the growth rate changes predicted by the Cholodny-Went theory.     

Blue and Green Light-Induced Phototropism in Arabidopsis thaliana and Lactuca sativa L. Seedlings

Exposure time-response curves for blue and green light-induced phototropic bending in hypocotyls of Arabidopsis thaliana (L.) Heynh. and Lactuca sativa L. seedlings are presented. These seedlings show significant phototropic sensitivity up to 540 to 550 nanometers. Since wave-lengths longer than 560 nanometers do not induce phototropic bending, it is suggested that the response to 510 to 550 nanometers light is mediated by the specific blue light photoreceptor of phototropism. We advise care in the use of green `safelights' for studies of phototropism.     

Phototropism of Arabidopsis thaliana in microgravity and fractional gravity on the International Space Station

While there is a great deal of knowledge regarding plant growth and development in microgravity aboard orbiting spacecraft, there is little information available about these parameters in reduced or fractional gravity conditions (less than the nominal 1g on Earth). Thus, in these experiments using the European Modular Cultivation System on the International Space Station, we studied the interaction between phototropism and gravitropism in the WT and mutants of phytochrome A and B of Arabidopis thaliana. Fractional gravity and the 1 g control were provided by centrifuges in the spaceflight hardware, and unidirectional red and blue illumination followed a white light growth period in the time line of the space experiments. The existence of red-light-based positive phototropism in hypocotyls of seedlings that is mediated by phytochrome was confirmed in these microgravity experiments. Fractional gravity studies showed an attenuation of red-light-based phototropism in both roots and hypocotyls of seedlings occurring due to gravitational accelerations ranging from 0.l to 0.3 g. In contrast, blue-light negative phototropism in roots, which was enhanced in microgravity compared with the 1g control, showed a significant attenuation at 0.3 g. In addition, our studies suggest that the well-known red-light enhancement of blue-light-induced phototropism in hypocotyls is likely due to an indirect effect by the attenuation of gravitropism. However, red-light enhancement of root blue-light-based phototropism may occur via a more direct effect on the phototropism system itself, most likely through the phytochrome photoreceptors. To our knowledge, these experiments represent the first to examine the behavior of flowering plants in fractional or reduced gravity conditions.     

Auxin Redistribution during First Positive Phototropism in Corn Coleoptiles : Microtubule Reorientation and the Cholodny-Went Theory

In red-light grown corn (Zea mays L. cv Brio42.HT) coleoptiles, cortical microtubules adjacent to the outer cell wall of the outer epidermis reorient from transverse to longitudinal in response to auxin depletion and after phototropic stimulation in the lighted side of the coleoptile. This was used as an in situ assay of cellular auxin concentration. The fluence-response relation for the blue light-induced reorientation is compared with that for first positive phototropism and the dose-response relationship for the auxin-dependent reorientation. The result supports the theory by Cholodny and Went, claiming that phototropic stimulation results in auxin displacement across the coleoptile. In terms of microtubule orientation, this displacement becomes even more pronounced after preirradiation with a weak blue light pulse from above.     

Phototropism and gravitropism in lateral roots of Arabidopsis

Gravitropism and, to a lesser extent, phototropism have been characterized in primary roots, but little is known about structural/functional aspects of these tropisms in lateral roots. Therefore, in this study, we report on tropistic responses in lateral roots of Arabidopsis thaliana. Lateral roots initially are plagiogravitropic, but when they reach a length of approximately 10 mm, these roots grow downward and exhibit positive orthogravitropism. Light and electron microscopic studies demonstrate a correlation between positive gravitropism and development of columella cells with large, sedimented amyloplasts in wild-type plants. Lateral roots display negative phototropism in response to white and blue light and positive phototropism in response to red light. As is the case with primary roots, the photoresponse is weak relative to the graviresponse, but phototropism is readily apparent in starchless mutant plants, which are impaired in gravitropism. To our knowledge, this is the first report of phototropism of lateral roots in any plant species.     

Positive Phototropism in the Thallus of Bryopsis plumosa

Positive phototropism in the thallus of the marine coenocyticgreen alga Bryopsis plumosa was investigated in terms of themode of bending, the photosensitive zone and the effectivenessspectrum. The bending occurred as a consequence of a differencein growth rate between the illuminated and the shaded sidesof the thallus. Elongation on the shaded side was stimulatedwhile that on the illuminated side was inhibited. However, theoverall elongation rate was barely affected. Illumination witha fine beam revealed that a zone from approximately 80 to 120µm below the tip was the most photosensitive. The effectivenessspectrum showed that blue light (<550 nm) was most effective,with light at 467 nm having maximal effectiveness. (Received November 29, 1994; Accepted May 29, 1995)     

A genetic framework for fruit patterning in Arabidopsis thaliana

In the model plant Arabidopsis thaliana, the establishment of organ polarity leads to the expression of FILAMENTOUS FLOWER (FIL) and YABBY3 (YAB3) on one side of an organ. One important question that has remained unanswered is how does this positional information lead to the correct spatial activation of genes controlling tissue identity? We provide the first functional link between polarity establishment and the regulation of tissue identity by showing that FIL and YAB3 control the non-overlapping expression patterns of FRUITFULL (FUL) and SHATTERPROOF (SHP), genes necessary to form stripes of valve margin tissue that allow the fruit to shatter along two defined borders and disperse the seeds. FIL and YAB3 activate FUL and SHP redundantly with JAGGED (JAG), a gene that also promotes growth in organs, indicating that several pathways converge to regulate these genes. These activities are negatively regulated by REPLUMLESS (RPL), which divides FIL/JAG activity, creating two distinct stripes of valve margin.     

Sensitivity to Abscisic Acid Modulates Positive Interactions between Arabidopsis thaliana Individuals

The ability of abscisic acid (ABA) to modulate positive interactions between Arabidopsis thaliana individuals under salinity stress was investigated using abi1-1 (insensitive to ABA), era1-2 (hypersensitive to ABA) mutant and wild type plants. The results showed that sensitivity to ABA affects relative interaction intensity (RII) between Arabidopsis thaliana individuals. The neighbor removal experiments also confirmed the role of phenotypic responses in linking plant-plant interactions and sensitivity to ABA. For abi1-1 mutants, the absolute value differences between neighbor removal and control of stem length, root length, leaf area, leaf thickness, flower density, above biomass/belowground biomass (A/U), photosynthetic rate, stomatal conductance, leaf water content and water-use efficiency were smaller than those of the wild type, while for era1-2 mutants, these absolute value differences were larger than those of the wild type. Thus, it is suggested that positive interactions between Arabidopsis thaliana individuals are at least partly modulated by different sensitivity to ABA through different physiological and phenotypic plasticity.     

Sensitivity to Abscisic Acid Modulates Positive Interactions between Arabidopsis thaliana Individuals

The ability of abscisic acid (ABA) to modulate positive interactions between Arabidopsis thaliana individuals under salinity stress was investigated using abi1-1 (insensitive to ABA), era1-2 (hypersen- sitive to ABA) mutant and wild type plants. The results showed that sensitivity to ABA affects relative interaction intensity (RII) between Arabidopsis thaliana individuals. The neighbor removal experiments also confirmed the role of phenotypic responses in linking plant-plant interactions and sensitivity to ...     

Functional bioinformatics for Arabidopsis thaliana

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Functional bioinformatics for Arabidopsis thaliana

MOTIVATION: The genome of Arabidopsis thaliana, which has the best understood plant genome, still has approximately one-third of its genes with no functional annotation at all from either MIPS or TAIR. We have applied our Data Mining Prediction (DMP) method to the problem of predicting the functional classes of these protein sequences. This method is based on using a hybrid machine-learning/data-mining method to identify patterns in the bioinformatic data about sequences that are predictive of function. We use data about sequence, predicted secondary structure, predicted structural domain, InterPro patterns, sequence similarity profile and expressions data. RESULTS: We predicted the functional class of a high percentage of the Arabidopsis genes with currently unknown function. These predictions are interpretable and have good test accuracies. We describe in detail seven of the rules produced.     

A method for measuring whole plant photosynthesis in Arabidopsis thaliana

Measurement of photosynthesis of intact leaves of Arabidopsis thaliana has been prohibitive due to the small leaf size and prostrate growth habit. Because of the widespread use of Arabidopsis for plant science research it is important to have a procedure for accurate, nondestructive measurement of its photosynthesis. We developed and tested a method for analysis of photosynthesis in whole plants of Arabidopsis. Net carbon assimilation and stomatal conductance were measured with an open gas exchange system and photosynthetic oxygen evolution was determined from chlorophyll fluorescence parameters. Individual plants were grown in 50 cubic centimeter tubes that were attached with an air tight seal to an enclosed gas exchange chamber for measurement of carbon dioxide and water exchange by the whole plant. Chlorophyll fluorescence from intact leaves was simultaneously measured with a pulse modulated fluorometer. Photosynthetic CO₂ assimilation and stomatal conductance rates were calculated with established gas exchange procedures and O₂ evolution was determined from chlorophyll fluorescence measurement of Photosystem II yield. Carbon assimilation and oxygen evolution in response to light intensity and ambient CO₂ concentration was measured and is presented here to demonstrate the potential use of this method for investigation of photosynthesis of Arabidopsis plants in controlled environment conditions.     

A puromycin-sensitive aminopeptidase is essential for meiosis in Arabidopsis thaliana

Puromycin-sensitive aminopeptidases (PSAs) participate in a variety of proteolytic events essential for cell growth and viability, and in fertility in a broad range of organisms. We have identified and characterized an Arabidopsis thaliana mutant (mpa1) from a pool of T-DNA tagged lines that lacks PSA activity. This line exhibits reduced fertility, producing shorter siliques (fruits) bearing a lower number of seeds compared with wild-type plants. Cytogenetic characterization of meiosis in the mutant line reveals that both male and female meiosis are defective. In mpa1, early prophase I appears normal, but after pachytene most of the homologous chromosomes are desynaptic, thus, by metaphase I a high level of univalence is observed subsequently leading to abnormal chromosome segregation. Wild-type plants treated with specific inhibitors of PSA show a very similar desynaptic phenotype to that of the mutant line. A fluorescent PSA-specific bioprobe, DAMPAQ-22, reveals that the protein is maximally expressed in wild-type meiocytes during prophase I and is absent in mpa1. Immunolocalization of meiotic proteins showed that the meiotic recombination pathway is disrupted in mpa1. Chromosome pairing and early recombination appears normal, but progression to later stages of recombination and complete synapsis of homologous chromosomes are blocked.     

A novel reporter for intrachromosomal homoeologous recombination in Arabidopsis thaliana

A reporter system using engineered introns as recombination substrates in the uidA (GUS) gene has been developed and characterized in Arabidopsis thaliana. The non-coding nature of the recombination substrate has allowed us to monitor recombination events between duplicated copies of the intron that are either identical (homologous recombination) or harbour sequence polymorphisms (homoeologous recombination). The effects of substrate length and divergence on the frequency of recombination events were examined. A positive correlation between substrate length and somatic recombination frequency was found as the frequency of recombination increased 183-fold when the recombination substrate was lengthened from 153 to 589 bp. The existence of 11 polymorphisms in a 589-bp recombination substrate (1.9% sequence divergence) led to an almost 10-fold reduction in the frequency of recombination. This result demonstrates that relatively modest levels of sequence divergence can substantially reduce the frequency of recombination in plants. A molecular analysis of recombination products revealed that the recombination junctions are more frequent in the central segment of the recombination substrate.     

A new seed-based assay for meiotic recombination in Arabidopsis thaliana

Meiotic recombination is a fundamental biological process that plays a central role in the evolution and breeding of plants. We have developed a new seed-based assay for meiotic recombination in Arabidopsis. The assay is based on the transformation of green and red fluorescent markers expressed under a seed-specific promoter. A total of 74 T-DNA markers were isolated, sequenced and mapped both physically and genetically. Lines containing red and green markers that map 1-20 cM apart were crossed to produce tester lines with the two markers linked in cis yielding seeds that fluoresced both in red and green. We show that these lines can be used for efficient scoring of recombinant types (red only or green only fluorescing seeds) in a seed population derived from a test cross (backcross) or self-pollination. Two tester lines that were characterized during several generations of backcross and self-pollination, one in the background of ecotype Landsberg and one in the ecotype Columbia, are described. We discuss the number of plants and seeds to be scored in order to obtain reliable and reproducible crossing over rate values. This assay offers a relatively high-throughput method, with the benefit of seed markers (similar to the maize classical genetic markers) combined with the advantages of Arabidopsis. It advances the prospect to better understand the factors that affect the rate of meiotic crossover in plants and to stimulate this process for more efficient breeding and mapping.     

A convenient and versatile hydroponic cultivation system for Arabidopsis thaliana

A versatile two-step cultivation procedure for Arabidopsis thaliana is described for the production of large quantities of leaf material suitable for biochemical and biophysical analysis. The first step comprises a miniature greenhouse made out of a plastic pipette box to grow the seedlings to the six-leaf stage. For continued growth, the seedlings are transferred to hydroponic cultivation using an opaque container covered by a styrofoam lid. Transfer of the small seedlings to hydroponic culture is facilitated by growth in separate pipette tips, which protects vulnerable roots from damage. The hydroponic cultivation system is easy to scale-up and produces large amounts of relatively large leaves and roots. This hydroponic system produces enough plant material to make Arabidopsis a feasible model for biochemical and biophysical experiments, which can be combined with the available genetic information to address various aspects of plant functional genomics.     

A novel method for growing Arabidopsis thaliana plants hydroponically

In this report we describe the development of a simple system for growing individual Arabidopsis plants hydroponically under environmental conditions which are simple to maintain while maximizing growth and development. The system consists of a Life Raft Float Unit with a 14–20 mm sponge inserted into its center; this unit is placed into a Magenta GA7 vessel containing Arabidopsis nutrient solution. Plants grown in this experimental system do not require aeration, 125 μmol m⁻² s⁻¹ supports vigorous plant growth, 1/4 to 1/8 strength Arabidopsis nutrient solution promotes maximal growth while higher or lower levels have adverse effects and temperatures between 18 and 24°C were found to be optimal. Plants grown under these conditions exhibit physiological factors such as time to flowering and overall appearance similar to those grown in potting media. The low maintenance method described in this paper provides an alternative to existing hydroponic methods used to grow Arabidopsis . Adaptation of this system for growing large numbers of plants in 1 container is also discussed.