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All sourcesNASA NTRSDOE OSTI
NASA NTRS1988

The habits of roots: what's up down under?

Defining interactions of roots with the surrounding soil environment has been the focus of many recent investigations. As a result of these efforts, we are gaining an appreciation of the varied and often surprising strategies whereby roots adjust to and condition their soil environment for optimal growth and development. This article summarizes current knowledge of the often complex interactions between roots and biotic and abiotic factors within the soil. These interactions are interpreted in terms of modifications in the development or the physiology of the root.

Review↗
DOE OSTI2021

The Landscape of RNA-Protein Interactions in Plants: Approaches and Current Status

RNAs transmit information from DNA to encode proteins that perform all cellular processes and regulate gene expression in multiple ways. From the time of synthesis to degradation, RNA molecules are associated with proteins called RNA-binding proteins (RBPs). The RBPs play diverse roles in many aspects of gene expression including pre-mRNA processing and post-transcriptional and translational regulation. In the last decade, the application of modern techniques to identify RNA–protein interactions with individual proteins, RNAs, and the whole transcriptome has led to the discovery of a hidden landscape of these interactions in plants. Global approaches such as RNA interactome capture (RIC) to identify proteins that bind protein-coding transcripts have led to the identification of close to 2000 putative RBPs in plants. Interestingly, many of these were found to be metabolic enzymes with no known canonical RNA-binding domains. Here, we review the methods used to analyze RNA–protein interactions in plants thus far and highlight the understanding of plant RNA–protein interactions these techniques have provided us. We also review some recent protein-centric, RNA-centric, and global approaches developed with non-plant systems and discuss their potential application to plants. We also provide an overview of results from classical studies of RNA–protein interaction in plants and discuss the significance of the increasingly evident ubiquity of RNA–protein interactions for the study of gene regulation and RNA biology in plants.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
NASA NTRS1987

Collection of gravitropic effectors from mucilage of electrotropically-stimulated roots of Zea mays L

We placed agar blocks adjacent to tips of electrotropically stimulated primary roots of Zea mays. Blocks placed adjacent to the anode-side of the roots for 3 h induced significant curvature when subsequently placed asymmetrically on tips of vertically-oriented roots. Curvature was always toward the side of the root unto which the agar block was placed. Agar blocks not contacting roots and blocks placed adjacent to the cathode-side of electrotropically stimulated roots did not induce significant curvature when placed asymmetrically on tips of vertically-oriented roots. Atomic absorption spectrophotometry indicated that blocks adjacent to the anode-side of electrotropically-stimulated roots contained significantly more calcium than (1) blocks not contacting roots, and (2) blocks contacting the cathode-side of roots. These results demonstrate the presence of a gradient of endogenous Ca in mucilage of electrotropically-stimulated roots (i.e. roots undergoing gravitropic-like curvature).

NASA Discipline Number 40-10↗
NASA NTRS1990

A direct screening procedure for gravitropism mutants in Arabidopsis thaliana (L.) Heynh

In order to isolate gravitropism mutants of Arabidopsis thaliana (L.) Heynh. var Estland for the genetic dissection of the gravitropism pathway, a direct screening procedure has been developed in which mutants are selected on the basis of their gravitropic response. Variability in hypocotyl curvature was dependent on the germination time of each seed stock, resulting in the incorrect identification of several lines as gravitropism mutants when a standard protocol for the potentiation of germination was used. When the protocol was adjusted to allow for differences in germination time, these lines were eliminated from the collection. Out of the 60,000 M2 seedlings screened, 0.3 to 0.4% exhibited altered gravitropism. In approximately 40% of these mutant lines, only gravitropism by the root or the hypocotyl was altered, while the response of the other organ was unaffected. These data support the hypothesis that root and hypocotyl gravitropism are genetically separable.

NASA Discipline Number 29-20↗
NASA NTRS1988

Kinetics of determination in the differentiation of isolated mesophyll cells of Zinnia elegans to tracheary elements

Mechanically isolated mesophyll cells of Zinnia elegans L. cv Envy differentiate to tracheary elements when cultured in inductive medium containing 0.5 micromolar alpha-naphthaleneacetic acid and 0.5 micromolar benzyladenine. The cells do not differentiate when cultured in medium in which the concentration of auxin and/or cytokinin has been reduced to 0.005 micromolar. Cells require an initial 24-hour exposure to inductive cytokinin and 56-hour exposure to inductive auxin for differentiation at 72 hours of culture. Freshly isolated Zinnia cells can be maintained in medium having low concentrations of both auxin and cytokinin for only 1 day without significant loss of potential to differentiate upon transfer to inductive medium. Initial culture for up to 2 days in medium having high auxin and low cytokinin, or low auxin and high cytokinin, allows full differentiation on the third day after transfer to inductive medium and potentiates the early differentiation of some cells.

Non-NASA Center↗
NASA NTRS1987

The locations and amounts of endogenous ions and elements in the cap and elongating zone of horizontally oriented roots of Zea mays L.: an electron-probe EDS study

We used quantitative electron-probe energy-dispersive x-ray microanalysis to localize endogenous Na, Cl, K, P, S, Mg and Ca in cryofixed and freeze-dried cryosections of the cap (i.e. the putative site of graviperception) and elongating zone (i.e. site of gravicurvature) of horizontally oriented roots of Zea mays. Ca, Na, Cl, K and Mg accumulate along the lower side of caps of horizontally oriented roots. The most dramatic asymmetries of these ions occur in the apoplast, especially the mucilage. We could not detect any significant differences in the concentrations of these ions in the central cytoplasm of columella cells along the upper and lower sides of caps of horizontally-oriented roots. However, the increased amounts of Na, Cl, K and Mg in the longitudinal walls of columella cells along the lower side of the cap suggest that these ions may move down through the columella tissue of horizontally-oriented roots. Ca also accumulates (largely in the mucilage) along the lower side of the elongating zone of horizontally-oriented roots, while Na, P, Cl and K tend to accumulate along the upper side of the elongating zone. Of these ions, only K increases in concentration in the cytoplasm and longitudinal walls of cortical cells in the upper vs lower sides of the elongating zone. These results indicate that (1) gravity-induced asymmetries of ions differ significantly in the cap and elongating zone of graviresponding roots, (2) Ca accumulates along the lower side of the cap and elongating zone of graviresponding roots, (3) increased growth of the upper side of the elongating zone of horizontally-oriented roots correlates positively with increased amounts of K in the cytoplasm and longitudinal walls of cortical cells, and (4) the apoplast (especially the mucilage) may be an important component of the pathway via which ions move in graviresponding rots of Zea mays. These results are discussed relative to mechanisms for graviperception and gravicurvature of roots.

NASA Program Space Biology↗
NASA NTRS1985

In vivo inhibition of polyamine biosynthesis and growth in tobacco ovary tissues

Post fertilization growth of tobacco ovary tissues treated with inhibitors of polyamine (PA) biosynthesis was examined in relation to endogenous PA titers and the activities of arginine decarboxylase (ADC, EC 4.1.1.19) and ornithine decarboxylase (ODC, EC 4.1.1.17). DL-alpha-Difluoromethylornithine (DFMO) and DL-alpha-difluoromethylarginine (DFMA), specific, irreversible ("suicide") inhibitors of ODC and ADC in vitro, were used to modulate PA biosynthesis in excised flowers. ODC represented >99% of the total decarboxylase activity in tobacco ovaries. In vivo inhibition of ODC with DFMO resulted in a significant decrease in PA titers, ovary fresh weight and protein content. Simultaneous inhibition of both decarboxylases by DFMO and DFMA produced only a marginally greater depression in growth and PA titers, indicating that ODC activity is rate-limiting for PA biosynthesis in these tissues. Paradoxically, DFMA alone inhibited PA biosynthesis, not as a result of a specific inhibition of ADC, but primarily through the inactivation of ODC. In vivo inhibition of ODC by DFMA appears to result from arginase-mediated hydrolysis of this inhibitor to urea and DFMO, the suicide substrate for ODC. Putrescine conjugates in tobacco appear to function as a storage form of this amine which, upon hydrolysis, may contribute to Put homeostasis during growth.

NASA Program Space Biology↗
NASA NTRS1990

Characterization and distribution of a maize cDNA encoding a peptide similar to the catalytic region of second messenger dependent protein kinases

Maize (Zea mays) roots respond to a variety of environmental stimuli which are perceived by a specialized group of cells, the root cap. We are studying the transduction of extracellular signals by roots, particularly the role of protein kinases. Protein phosphorylation by kinases is an important step in many eukaryotic signal transduction pathways. As a first phase of this research we have isolated a cDNA encoding a maize protein similar to fungal and animal protein kinases known to be involved in the transduction of extracellular signals. The deduced sequence of this cDNA encodes a polypeptide containing amino acids corresponding to 33 out of 34 invariant or nearly invariant sequence features characteristic of protein kinase catalytic domains. The maize cDNA gene product is more closely related to the branch of serine/threonine protein kinase catalytic domains composed of the cyclic-nucleotide- and calcium-phospholipid-dependent subfamilies than to other protein kinases. Sequence identity is 35% or more between the deduced maize polypeptide and all members of this branch. The high structural similarity strongly suggests that catalytic activity of the encoded maize protein kinase may be regulated by second messengers, like that of all members of this branch whose regulation has been characterized. Northern hybridization with the maize cDNA clone shows a single 2400 base transcript at roughly similar levels in maize coleoptiles, root meristems, and the zone of root elongation, but the transcript is less abundant in mature leaves. In situ hybridization confirms the presence of the transcript in all regions of primary maize root tissue.

NASA Program Space Biology↗
NASA NTRS1988

Hormonal induction and antihormonal inhibition of tracheary element differentiation in Zinnia cell cultures

Mechanically isolated mesophyll cells of Zinnia elegans L. cv Envy differentiate to tracheary elements when cultured in inductive medium containing sufficient auxin and cytokinin. Tracheary element differentiation was induced by the three auxins (alpha-naphthaleneacetic acid, indole-3-acetic acid, and 2,4-dichlorophenoxyacetic acid) and four cytokinins (6-benzyladenine, kinetin, 2-isopentenyladenine and zeatin) tested. Tracheary element formation is inhibited or delayed if the inductive medium is supplemented with an anticytokinin, antiauxin, or inhibitor of auxin transport.

NASA Program Space Biology↗
NASA NTRS1986

Amyloplast sedimentation and organelle saltation in living corn columella cells

Amyloplast sedimentation during gravistimulation and organelle movements was studied in living central rootcap cells of Zea mays L. cv. Merit. Cells from sectioned roots were viewed with a horizontally-mounted videomicroscope. The kinetics of gravity-induced amyloplast sedimentation were comparable to those calculated from experiments using fixed material. Individual amyloplasts fell at an average velocity of 5.5 micrometers min-1; the maximal velocity of fall measured was 18.0 micrometers min-1. Amyloplasts often rotated, sometimes rose in the cytoplasm, and occasionally underwent sudden rapid movements as fast as 58 micrometers min-1. Saltations of other organelles were frequently observed. This appears to be the first report of cytoplasmic streaming in the presumptive statocytes of roots.

NASA Program Space Biology↗
NASA NTRS1989

Specific photoaffinity labeling of two plasma membrane polypeptides with an azido auxin

Plasma membrane vesicles were isolated from zucchini (Cucurbita pepo) hypocotyl tissue by aqueous phase partitioning and assessed for homogeneity by the use of membrane-specific enzyme assays. The highly pure (ca. 95%) plasma membrane vesicles maintained a pH differential across the membrane and accumulated a tritiated azido analogue of 3-indoleacetic acid (IAA), 5-azido-[7-3H]IAA ([3H]N3IAA), in a manner similar to the accumulation of [3H]IAA. The association of the [3H]N3IAA with membrane vesicles was saturable and subject to competition by IAA and auxin analogues. Auxin-binding proteins were photoaffinity labeled by addition of [3H]N3IAA to plasma membrane vesicles prior to exposure to UV light (15 sec; 300 nm) and detected by subsequent NaDodSO4/PAGE and fluorography. When the reaction temperature was lowered to -196 degrees C, high-specific-activity labeling of a 40-kDa and a 42-kDa polypeptide was observed. Triton X-100 (0.1%) increased the specific activity of labeling and reduced the background, which suggests that the labeled polypeptides are intrinsic membrane proteins. The labeled polypeptides are of low abundance, as expected for auxin receptors. Further, the addition of IAA and auxin analogues to the photoaffinity reaction mixture resulted in reduced labeling that was qualitatively similar to their effects on the accumulation of radiolabeled IAA in membrane vesicles. Collectively, these results suggest that the radiolabeled polypeptides are auxin receptors. The covalent nature of the label should facilitate purification and further characterization of the receptors.

NASA Discipline Number 29-20↗
NASA NTRS1992

Clinostats and Centrifuges: Their Use, Value, and Limitations in Gravitational Biological Research

The present volume addresses physical phenomena and effects associated with clinostat and centrifuge operations as well as their physiological effects. Particular attention is given to the simulation of the gravity conditions on the ground, the internal dynamics of slowly rotating biological systems, and qualitative and quantitative aspects of the fast-rotating clinostat as a research tool. Also discussed are the development and use of centrifuges in gravitational biology, the use of centrifuges in plant gravitational biology and a comparison of ground-based and flight experiment results, the ability of clinostat to mimic the effect of microgravity on plant cells and organs, and the impact of altered gravity conditions on early EGF-induced signal transduction in human epidermal A431 cells.

Halstead, Thora W.↗
NASA NTRS1987

Occurrence and metabolism of 7-hydroxy-2-indolinone-3-acetic acid in Zea mays

7-Hydroxy-2-indolinone-3-acetic acid was identified as a catabolite of indole-3-acetic acid in germinating kernels of Zea mays and found to be present in amounts of ca 3.1 nmol/kernel. 7-Hydroxy-2-indolinone-3-acetic acid was shown to be a biosynthetic intermediate between 2-indolinone-3-acetic acid and 7-hydroxy-2-indolinone-3-acetic acid-7'-O-glucoside in both kernels and roots of Zea mays. Further metabolism of 7-hydroxy-2-[5-3H]-indolinone-3-acetic acid-7'-O-glucoside occurred to yield tritiated water plus, as yet, uncharacterized products.

NASA Program Space Biology↗
NASA NTRS1991

Hydrotropism and its interaction with gravitropism in maize roots

We have partially characterized root hydrotropism and its interaction with gravitropism in maize (Zea mays L.). Roots of Golden Cross Bantam 70, which require light for orthogravitropism, showed positive hydrotropism; bending upward when placed horizontally below a hydrostimulant (moist cheesecloth) in 85% relative humidity (RH) and in total darkness. However, the light-exposed roots of Golden Cross Bantam 70 or roots of a normal maize cultivar, Burpee Snow Cross, showed positive gravitropism under the same conditions; bending downward when placed horizontally below the hydrostimulant in 85% RH. Light-exposed roots of Golden Cross Bantam 70 placed at 70 degrees below the horizontal plane responded positively hydrotropically, but gravitropism overcame the hydrotropism when the roots were placed at 45 degrees below the horizontal. Roots placed vertically with the tip down in 85% RH bent to the side toward the hydrostimulant in both cultivars, and light conditions did not affect the response. Such vertical roots did not respond when the humidity was maintained near saturation. These results suggest that hydrotropic and gravitropic responses interact with one another depending on the intensity of one or both factors. Removal of the approximately 1.5 millimeter root tip blocked both hydrotropic and gravitropic responses in the two cultivars. However, removal of visible root tip mucilage did not affect hydrotropism or gravitropism in either cultivar.

NASA Program Space Biology↗
NASA NTRS1990

Temperature sensing by primary roots of maize

Zea mays L. seedlings, grown on agar plates at 26 degrees C, reoriented the original vertical direction of their primary root when exposed to a thermal gradient applied perpendicular to the gravity vector. The magnitude and direction of curvature can not be explained simply by either a temperature or a humidity effect on root elongation. It is concluded that primary roots of maize sense temperature gradients in addition to sensing the gravitational force.

NASA Program Space Biology↗
NASA NTRS2000

Interaction of root gravitropism and phototropism in Arabidopsis wild-type and starchless mutants

Root gravitropism in wild-type Arabidopsis and in two starchless mutants, pgm1-1 and adg1-1, was evaluated as a function of light position to determine the relative strengths of negative phototropism and of gravitropism and how much phototropism affects gravitropic measurements. Gravitropism was stronger than phototropism in some but not all light positions in wild-type roots grown for an extended period, indicating that the relationship between the two tropisms is more complex than previously reported. Root phototropism significantly influenced the time course of gravitropic curvature and the two measures of sensitivity. Light from above during horizontal exposure overestimated all three parameters for all three genotypes except the wild-type perception time. At the irradiance used (80 micromol m(-2) s(-1)), the shortest periods of illumination found to exaggerate gravitropism were 45 min of continuous illumination and 2-min doses of intermittent illumination. By growing roots in circumlateral light or by gravistimulating in the dark, corrected values were obtained for each gravitropic parameter. Roots of both starchless mutants were determined to be about three times less sensitive than prior estimates. This study demonstrates the importance of accounting for phototropism in the design of root gravitropism experiments in Arabidopsis.

Non-NASA Center↗
NASA NTRS1988

Light-regulated protein and mRNA synthesis in root caps of maize

Illumination of maize roots initiates changes in mRNA levels and in the activities of proteins within the root cap. Using Northern analysis we showed a 5-6 fold increase in the levels of three specific mRNAs and a 14-fold increase in plastid mRNA. This increase is rapid, occurring within 30 minutes of illumination. With prolonged periods of darkness following illumination, messages return to levels observed in dark, control caps. For two species of mRNA illumination results in a reduction in message levels. Light-stimulated increases in the levels of specific mRNAs are proportionally greater than are increases in the activities of corresponding proteins. We suggest that the light-stimulated increase in protein activity in root caps may be preceded by and occur as a consequence of enhanced levels of mRNA. Our work suggests that photomorphogenesis in roots could involve changes in the levels of a wide variety of mRNAs within the root cap.

NASA Program Space Biology↗
NASA NTRS1987

Shoot inversion inhibition of stem elongation in Pharbitis nil: a possible role for ethylene-induced glycoprotein and lignin

Inversion of the upper shoot of Pharbitis nil results in the inhibition of elongation in the inverted stem. The objective of the present study was to determine how shoot inversion-induced gravity stress inhibited elongation and to elucidate the possible role of ethylene-induced glycoprotein and lignin in this process. Determinations of hydroxyproline, peroxidase, phenylalanine ammonia-lyase (PAL), phenol, and lignin content/activity were carried out by appropriate spectrophotometric methods. It was found that inversion and Ethrel treatments of upright shoots caused significant increases in hydroxyproline content, peroxidase, and PAL activity in 12 hours and in phenol and lignin contents in 24 hours. All of these increases except for that of cytoplasmic peroxidase activity were partially reversed by AgNO3, the ethylene action inhibitor. It is concluded that possible cross-linking associated with the accumulation of the ethylene-induced hydroxyproline-rich glycoprotein and lignin may be responsible for the later stages of cessation of elongation in the inverted Pharbitis shoot.

NASA Program Space Biology↗