Fred Rook

Sugar Sensing and Sugar-Mediated Signal Transduction in Plants

Information concerning the sugar status of plant cells is of great importance during all stages of the plant life cycle. The availability of or lack of sugars triggers many metabolic and developmental responses, and it is not surprising, therefore, that sugars profoundly affect the expression of a large number of genes (for review, see Koch, 1996; Graham, 1996). Sugar sensing occurs at the level of individual cells and the responses of such cells must be integrated at the tissue, organ, and plant level. Therefore, sugar-induced signals will interact with other sensing and signaling pathways. The mechanisms used by plant cells to sense sugars and to process this information are essentially unknown, and only recently are these questions being addressed experimentally. This lack of knowledge contrasts with the situation in yeast and bacteria, in which the molecular and physiological analysis of mutants have yielded extensive information about sugar perception (Trumbly, 1992; Ronne, 1995; Saier et al., 1995).

Characterization of Mutants in Arabidopsis Showing Increased Sugar-Specific Gene Expression, Growth, and Developmental Responses

Sugars such as sucrose serve dual functions as transported carbohydrates in vascular plants and as signal molecules that regulate gene expression and plant development. Sugar-mediated signals indicate carbohydrate availability and regulate metabolism by co-coordinating sugar production and mobilization with sugar usage and storage. Analysis of mutants with altered responses to sucrose and glucose has shown that signaling pathways mediated by sugars and abscisic acid interact to regulate seedling development and gene expression. Using a novel screen for sugar-response mutants based on the activity of a luciferase reporter gene under the control of the sugar-inducible promoter of the ApL3 gene, we have isolated high sugar-response (hsr) mutants that exhibit elevated luciferase activity and ApL3 expression in response to low sugar concentrations. Our characterization of these hsr mutants suggests that they affect the regulation of sugar-induced and sugar-repressed processes controlling gene expression, growth, and development in Arabidopsis. In contrast to some other sugar-response mutants, they do not exhibit altered responses to ethylene or abscisic acid, suggesting that the hsr mutants may have a specifically increased sensitivity to sugars. Further characterization of the hsr mutants will lead to greater understanding of regulatory pathways involved in metabolite signaling.

Methotrexate does not block import of a DHFR fusion protein into chloroplasts

Protein import into chloroplasts requires the movement of a precursor protein across the envelope membranes. The conformation of a precursor as it passes from the aqueous medium across the hydrophobic membranes is not known in detail. To address this problem we examined precursor conformation during translocation using the chimeric precursor PCDHFR, which contains the plastocyanin (PC) transit peptide in front of mouse cytosolic dihydrofolate reductase (DHFR). The chimeric protein is targeted to chloroplasts and is competent for import. The conformation of PCDHFR can be stabilized by complexing with methotrexate, an analogue of the substrate of DHFR. Methotrexate strongly inhibits DHFR import into yeast mitochondria (M. Eilers and G. Schatz, Nature 322 (1986) 228–232), presumably because the precursor must unfold to cross the membrane and it cannot do so when complexed with methotrexate. We show here that methotrexate does not block PCDHFR import into chloroplasts. Methotrexate does slow the rate of import, and protects DHFR from degradation once inside chloroplasts. The processed protein is localized in the stroma, indicating that import into thylakoids is impeded. Protease sensitivity assays indicate that the complex of precursor protein with methotrexate changes in conformation during the translocation across the envelope.

The light-regulated Arabidopsis bZIP transcription factor gene ATB2 encodes a protein with an unusually long leucine zipper domain

A light-regulated basic domain/leucine zipper gene, ATB2, was identified in an Arabidopsis thaliana transcription factor gene collection. Both genomic and cDNA clones of ATB2 were isolated. The gene encodes a small protein (18 kDa) which mainly consists of the basic domain and an unusually long leucine zipper. The expression of the ATB2 gene is induced when etiolated or dark-adapted seedlings are transferred to the light. Moreover, its expression is derepressed in dark-grown seedlings of the photomorphogenic mutants cop1 and det1. In mature plants, transcript levels are particularly high in flowers and also light-responsive in these tissues.

Carbohydrate Status of Tulip Bulbs during Cold-Induced Flower Stalk Elongation and Flowering

The effect of a cold treatment on the carbohydrate status of the scales and flower stalk of Tulipa gesneriana L. cv Apeldoorn bulbs during growth after planting was studied and compared with bulbs not given cold treatment. Bulbs were stored dry for 12 weeks at 5[deg]C (precooled) or 17[deg]C (noncooled). Only the 5[deg]C treatment led to rapid flower stalk elongation and flowering following planting at higher temperatures. Precooling enhanced mobilization of starch, fructans, and sucrose in the scales. The cold-stimulated starch breakdown was initially accompanied by increased [alpha]-amylase activity per scale. In noncooled bulbs, [alpha]-amylase activity slightly decreased or remained more or less constant. Cold-induced flower stalk elongation was partially accompanied by a decrease in the sucrose content and an increase in the glucose content and invertase activity per g dry weight. The starch content in internodes initially decreased and subsequently increased; [alpha]-amylase activity per g dry weight of the lowermost internode showed a peak pattern during starch breakdown and increased thereafter. The internodes of noncooled bulbs, on the contrary, accumulated sucrose. Their glucose content and invertase activity per g dry weight remained low. Starch breakdown was not found and [alpha]-amylase activity per g dry weight of the lowermost internode remained at a low level. Precooling of tulip bulbs thus favors reserve mobilization in the scales and flower stalk and glucose accumulation in the elongating internodes.

Do Transcription Factors Play Special Roles in Adaptive Variation

Ever since the work by Jacob and Monod on the Lac operon, scientists have appreciated that the control of gene expression is one of the most important points of regulation in biology. Although many other layers of regulation exist, the possibility of control at the start point of production of RNA

Sucrose is a signalling molecule in plants

Because of their sessile nature plants must adapt to their environment. Plants receive signals and react to them by changing their metabolic state and/or development. Light is a very important environmental signal. Differences in light intensity, duration of the light period and composition of the spectrum have profound influence on plant growth and development. This is already demonstrated in young seedlings. When growing in darkness seedlings develop an elongated hypocotyl and closed yellow cotyledons. When seedlings are put in the light hypocotyl elongation is inhibited and the cotyledons expand and turn green. Light is perceived by photoreceptors of which the phytochromes are the best studied. Next the light signal is transduced involving a signalling cascade including G-proteins, Ca2+, Calmodulin and cGMP eventually leading to changes in gene expression and developmental program [1]. In Arabidopsis several mutants in light dependent development were isolated. The hy (long-hypocotyl) mutants show dark grown aspects when grown in the light [2] and the groups of det (de-etiolated) and cop (constitutive photomorphogenic) mutants [3] show light grown aspects when grown in the dark. Changes in gene expression in response to light involve transcription factors. Transcription factors are DNA binding proteins that bind to the promoter region of their target gene and induce or repress gene expression.

[12] Analysis of light-regulated gene expression

Publisher Summary Light is important for plant development and the expression of a large number of genes is regulated by light. The quality, amount, and duration of the light treatment needed to induce or repress the expression of a gene of interest provide information about the way light controls its expression. The phytochromes are the best studied family of photoreceptors. Phytochrome can function as a molecular switch because many responses that are triggered by a pulse of red light can be reduced when the plant is subsequently exposed to far-red light. To determine the role of phytochrome in the light-regulated expression of the gene of interest, red and far-red light source are required. A red light source can be obtained by filtering the red light portion from an incandescent bulb with filters such as those used in photography. Moreover, several types of red-light bulbs and fluorescent tubes are also commercially available and their use is much preferred.