Quantitative analysis of statolith sedimentation behavior was accomplished using videomicroscopy of living columella cells of corn (Zea mays) roots, which displayed no systematic cytoplasmic streaming. Following 90°rotation of the root, the statoliths moved downward along the distal wall and then spread out along the bottom with an average velocity of 1.7 m min Ϫ1 . When statolith trajectories traversed the complete width or length of the cell, they initially moved horizontally toward channel-initiation sites and then moved vertically through the channels to the lower side of the reoriented cell where they again dispersed. These statoliths exhibited a significantly lower average velocity than those sedimenting on distal-toside trajectories. In addition, although statoliths undergoing distal-to-side sedimentation began at their highest velocity and slowed monotonically as they approached the lower cell membrane, statoliths crossing the cell's central region remained slow initially and accelerated to maximum speed once they reached a channel. The statoliths accelerated sooner, and the channeling effect was less pronounced in roots treated with cytochalasin D. Parallel ultrastructural studies of high-pressure frozen-freeze-substituted columella cells suggest that the low-resistance statolith pathway in the cell periphery corresponds to the sharp interface between the endoplasmic reticulum (ER)-rich cortical and the ER-devoid central region of these cells. The central region is also shown to contain an actin-based cytoskeletal network in which the individual, straight, actin-like filaments are randomly distributed. To explain these findings as well as the results of physical simulation experiments, we have formulated a new, tensegrity-based model of gravity sensing in columella cells. This model envisages the cytoplasm as pervaded by an actin-based cytoskeletal network that is denser in the ER-devoid central region than in the ER-rich cell cortex and is linked to stretch receptors in the plasma membrane. Sedimenting statoliths are postulated to produce a directional signal by locally disrupting the network and thereby altering the balance of forces acting on the receptors in different plasma membrane regions.For nearly 100 years the dense, starch-filled amyloplasts within the columella cells of the higher plant root cap have been proposed to serve as the gravitysensing structures of the plant root gravitropic system (Haberlandt, 1900;Nemec, 1900; Darwin, 1903). The sedimentation of these amyloplasts (or statoliths) in response to gravity, and the columella cells (or statocytes) in which they are found, have repeatedly been shown to be necessary for a proper root response to gravity (

No abstract

We have investigated the structural events associated with vacuole biogenesis in root tip cells of tobacco (Nicotiana tabacum) seedlings preserved by high-pressure freezing and freeze-substitution techniques. Our micrographs demonstrate that the lytic vacuoles (LVs) of root tip cells are derived from protein storage vacuoles (PSVs) by cell type-specific sets of transformation events. Analysis of the vacuole transformation pathways has been aided by the phytin-dependent black osmium staining of PSV luminal contents. In epidermal and outer cortex cells, the central LVs are formed by a process involving PSV fusion, storage protein degradation, and the gradual replacement of the PSV marker protein a-tonoplast intrinsic protein (TIP) with the LV marker protein g-TIP. In contrast, in the inner cortex and vascular cylinder cells, the transformation events are more complex. During mobilization of the stored molecules, the PSV membranes collapse osmotically upon themselves, thereby squeezing the vacuolar contents into the remaining bulging vacuolar regions. The collapsed PSV membranes then differentiate into two domains: (1) vacuole "reinflation" domains that produce pre-LVs, and (2) multilamellar autophagosomal domains that are later engulfed by the pre-LVs. The multilamellar autophagosomal domains appear to originate from concentric sheets of PSV membranes that create compartments within which the cytoplasm begins to break down. Engulfment of the multilamellar autophagic vacuoles by the pre-LVs gives rise to the mature LVs. During pre-LV formation, the PSV marker a-TIP disappears and is replaced by the LV marker g-TIP. These findings demonstrate that the central LVs of root cells arise from PSVs via cell type-specific transformation pathways.

Callus cells of Arabidopsis thaliana (cv. Landsberg erecta) were exposed for 8 h to a horizontal clinostat rotation (H, simulated weightlessness), a vertical clinostat rotation (V, clinostat control), or a stationary control (S) growth condition. The amount of glucose and fructose apparently decreased, while starch content increased in the H compared with the V- and S-treated cells. In order to investigate the influences of clinostat rotation on the cellular proteome further, the proteome alterations induced by horizontal and vertical clinostat rotation have been comparatively analysed by high-resolution two-dimensional (2-D) gel electrophoresis and mass spectrometry. Image analysis of silver-stained 2-D gels revealed that 80 protein spots showed quantitative and qualitative variations that were significantly (P <0.01) and reproducibly different between the clinorotated (H or V) and the stationary control samples. Protein spots excised from 2-D gels were analysed by microbe high performance liquid chromatography-ion trap-mass spectrometry (LC-IT-MS) to obtain the tandem mass (MS/MS) spectra. 18 protein spots, which showed significant expression alteration only under the H condition compared with those under V and S conditions, were identified. Of these proteins, seven were involved in stress responses, and four protein spots were identified as key enzymes in carbohydrate metabolism and lipid biosynthesis. Two reversibly glycosylated cell wall proteins were down-regulated in the H samples. Other proteins such as protein disulphide isomerase, transcription initiation factor IIF, and two ribosomal proteins also exhibited altered expression under the H condition. The data presented in this study illustrate that clinostat rotation of Arabidopsis callus cells has a significant impact on the expression of proteins involved in general stress responses, metabolic pathways, gene activation/transcription, protein synthesis, and cell wall biosynthesis.

No abstract

Exposure of Arabidopsis callus to microgravity has a significant impact on the expression of proteins involved in stress responses, carbohydrate metabolism, protein synthesis, intracellular trafficking, signaling, and cell wall biosynthesis. Microgravity is among the main environmental stress factors that affect plant growth and development in space. Understanding how plants acclimate to space microgravity is important to develop bioregenerative life-support systems for long-term space missions. To evaluate the spaceflight-associated stress and identify molecular events important for acquired microgravity tolerance, we compared proteomic profiles of Arabidopsis thaliana callus grown under microgravity on board the Chinese spacecraft SZ-8 with callus grown under 1g centrifugation (1g control) in space. Alterations in the proteome induced by microgravity were analyzed by high performance liquid chromatography-electrospray ionization-tandem mass spectrometry with isobaric tags for relative and absolute quantitation labeling. Forty-five proteins showed significant (p < 0.05) and reproducible quantitative differences in expression between the microgravity and 1g control conditions. Of these proteins, the expression level of 24 proteins was significantly up-regulated and that of 21 proteins was significantly down-regulated. The functions of these proteins were involved in a wide range of cellular processes, including general stress responses, carbohydrate metabolism, protein synthesis/degradation, intracellular trafficking/transportation, signaling, and cell wall biosynthesis. Several proteins not previously known to be involved in the response to microgravity or gravitational stimuli, such as pathogenesis-related thaumatin-like protein, leucine-rich repeat extension-like protein, and temperature-induce lipocalin, were significantly up- or down-regulated by microgravity. The results imply that either the normal gravity-response signaling is affected by microgravity exposure or that microgravity might inappropriately induce altered responses to other environmental stresses.

SUMMARYThe KNOTTED1 homeobox (KNOX) family transcription factors are essential for stem cell establishment and maintenance and regulate various aspects of development in all green plants. Expression patterns of the KNOX genes in the roots of plants have been reported, but their role in development remains unclear. Here we show how the KNAT1 gene is specifically involved in root skewing in Arabidopsis. The roots of two mutant alleles of KNAT1 (bp-1 and bp-5) exhibited exaggerated skewing to the right of gravity when grown on both vertical and tilted agar medium surfaces. This skewing phenotype was enhanced by treatments with low concentrations of propyzamide, and required auxin transport. The KNAT1 mutation substantially decreased basipetal auxin transport and increased auxin accumulation in the roots. Using a PIN2-GFP reporter and western blot analysis, we found that this alteration in auxin transport was accompanied by a decrease in PIN2 levels in the root tip. Decreased PIN2 expression in the mutant roots was not accompanied by reduced mRNA levels, suggesting that the KNAT1 mutations affected PIN2 expression at the posttranscriptional level. In vivo visualization of intracellular vacuolar targeting indicated that vacuolar degradation of PIN2-GFP was significantly promoted in the root tips of the bp allelic mutants. Together, these results demonstrate that KNAT1 negatively modulates root skewing, possibly by regulating auxin transport.

;Glycosyltransferases are enzymes that catalyze the attachment of a sugar molecule to specific acceptor molecules. These enzymes have been shown to play important roles in a number of biological processes. Whereas a large number of putative glycosyltransferase genes have been identified by genomic sequencing, the functions of most of these genes are unknown. Here we report the characterization of an Arabidopsis mutant, designated gaolaozhuangren1 (glz1), which is allelic to parvus characterized recently. The glz1 mutant exhibited a reduced plant stature, reduced size of organs in the shoot and dark-green leaves, indicating an important role of GLZ1 gene in normal development. The earliest GLZ1 expression appears at the shoot apical region of 4-d-old seedlings, which coincides with the onset of the glz1 morphological phenotypes. GLZ1 is expressed in a tissue-specific and developmentally regulated manner, predominantly in the stem and silique, and moderately in the flower. GLZ1 expression is strong in the midrib of rosette and cauline leaves; however, its expression was not detectable in the midrib of the cotyledon. Further analyses revealed that carbohydrate composition and distribution were aberrant in the glz1 mutant. These, together with the GLZ1 expression pattern, suggest a requirement for the GLZ1 function in normal sink-source transition during plant development.