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Plants exhibit an ultimate case of the intracellular motility involving rapid organelle trafficking and continuous streaming of the endoplasmic reticulum (ER). Although it was long assumed that the ER dynamics is actomyosin-driven, the responsible myosins were not identified, and the ER streaming was not characterized quantitatively. Here we developed software to generate a detailed velocity-distribution map for the GFP-labeled ER. This map revealed that the ER in the most peripheral plane was relatively static, whereas the ER in the inner plane was rapidly streaming with the velocities of up to ~3.5 [micro]m/sec. Similar patterns were observed when the cytosolic GFP was used to evaluate the cytoplasmic streaming. Using gene knockouts, we demonstrate that the ER dynamics is driven primarily by the ER-associated myosin XI-K, a member of a plant-specific myosin class XI. Furthermore, we show that the myosin XI deficiency affects organization of the ER network and orientation of the actin filament bundles. Collectively, our findings suggest a model whereby dynamic three-way interactions between ER, F-actin, and myosins determine the architecture and movement patterns of the ER strands, and cause cytosol hauling traditionally defined as cytoplasmic streaming. myosin XI | actin filament | cytoplasmic streaming | velocity distribution map | Arabidopsis thaliana doi/ 10.1073/pnas.0911482107

The plant hormone auxin plays a crucial role in regulating plant development and plant architecture. The directional auxin distribution within tissues depends on PIN transporters that are polarly localized on the plasma membrane. The PIN polarity and the resulting auxin flow directionality are mediated by the antagonistic actions of PINOID kinase and protein phosphatase 2A. However, the contribution of the PIN phosphorylation to the polar PIN sorting is still unclear. Here, we identified an evolutionarily conserved phosphorylation site within the central hydrophilic loop of PIN proteins that is important for the apical and basal polar PIN Iocalizations. Inactivation of the phosphorylation site in PIN1(Ala) resulted in a predominantly basal targeting and increased the auxin flow to the root tip. In contrast, the outcome of the phosphomimic PIN1(Asp) manipulation was a constitutive, PINOID-independent apical targeting of PIN1 and an increased auxin flow in the opposite direction. Furthermore, the PINl(Asp) functionally replaced PIN2 in its endogenous expression domain, revealing that the phosphorylation-dependent polarity regulation contributes to functional diversification within the PIN family. Our data suggest that PINOID-independent PIN phosphorylation at one single site is adequate to change the PIN polarity and, consequently, to redirect auxin fluxes between cells and provide the conceptual possibility and means to manipulate auxin-dependent plant development and architecture. cell polarity | auxin distribution | plant architecture www.pnas.org/cgi/doi/10.1073/pnas.0909460107

Multicellular organs are composed of distinct cell types with unique assemblages of translated mRNAs. Here, ribosome-associated mRNAs were immunopurified from specific cell populations of intact seedlings using Arabidopsis thaliana lines expressing a FLAG-epitope tagged ribosomal protein L18 (FLAG-RPL18) via developmentally regulated promoters. The profiling of mRNAs in ribosome complexes, referred to as the translatome, identified differentially expressed mRNAs in 21 cell populations defined by cell-specific expression of FLAG-RPL18. Phloem companion ceils of the root and shoot had the most distinctive translatomes. When seedlings were exposed to a brief period of hypoxia, a pronounced reprioritization of mRNA enrichment in the cell-specific translatomes occurred, including a ubiquitous rise in 49 mRNAs encoding transcription factors, signaling proteins, anaerobic metabolism enzymes, and uncharacterized proteins. Translatome profiling also exposed an intricate molecular signature of transcription factor (TF) family member mRNAs that was markedly reconfigured by hypoxia at global and cell-specific levels. In addition to the demonstration of the complexity and plasticity of cell-specific populations of ribosome-associated mRNAs, this study provides an in silico dataset for recognition of differentially expressed genes at the cell-, region-, and organ-specific levels. www.pnas.org/cgi/doi/10.1073/pnas.0906131106

Formins are present in all eukaryotes and are essential for the creation of actin-based structures responsible for diverse cellular processes. Because multicellular organisms contain large formin gene families, establishing the physiological functions of formin isoforms has been difficult. Using RNAi, we analyzed the function of all 9 formin genes within the moss Physcomitrella patens. We show that plants lacking class II formins (For2) are severely stunted and composed of spherical cells with disrupted actin organization. In contrast, silencing of all other formins results in normal elongated cell morphology and actin organization. Consistent with a role in polarized growth, For2 are apically localized in growing cells. We show that an N-terminal phosphatase tensin (PTEN)-like domain mediates apical localization. The PTEN-like domain is followed by a conserved formin homology (FH)1-FH2 domain, known to promote actin polymerization. To determine whether apical localization of any FH1-FH2 domain mediates polarized growth, we performed domain swapping. We found that only the class II FH1-FH2, in combination with the PTEN-like domain, rescues polarized growth, because it cannot be replaced with a similar domain from a For1. We used in vitro polymerization assays to dissect the functional differences between these FH1-FH2 domains. We found that both the FH1 and the FH2 domains from For2 are required to mediate exceptionally rapid rates of actin filament elongation, much faster than any other known formin. Thus, our data demonstrate that rapid rates of actin elongation are critical for driving the formation of apical filamentous actin necessary for polarized growth. Physcomitrella patens | moss | profilin | tip growth | RNAi

Problem statement: In this study, we considered transport corridors and sampling aspects to be the major indicators of ecological effects on Alder (Alnus subcordata) cambium cells dynamics. Approach: Thus, 240 cores were taken from forest-facing and road-facing trunk of Alder trees along Amre, Neka and Darab Kola transport corridors in hyrcanian Forests of Iran. Results showed that the roads corridor had significant effects on Alnus subcordata annual rings (p = 0.04) and bark growth (p Key words: Alnus subcordata, cambium cells, annual rings, bark, transport corridor, INTRODUCTION Transport corridors are becoming a focus of ecological research because of their distinctive structure, function and impact on surrounding ecosystems (12). Forman and Alexander (8) defined road corridors as [...]

The sparse inflorescence1 (spi1), Barren inflorescencel (Bif1), barren inflorescence2 (bif2), and barren stalk1 (ba1) mutants produce fewer branches and spikelets in the inflorescence due to defects in auxin biosynthesis, transport, or response. We report that spi1, bif1, and ba1, but not bif2, also function in promoting cell elongation in the inflorescence. DOI: 10.1534/genetics.108.099390

The occurrence of nonarticulated laticifers has been recorded in the literature for Apocynaceae, but little information is available on their structure and the chemical compounds in their latex. The objective of this study was to describe, anatomically and histochemically, the laticifers and latex found in the roots, stems, and leaves of Mandevilla atroviolacea (Stadelm) Woodson, a genus in the Apocynaceae. Laticifer anatomy was examined by light microscopy, and the chemical nature of the exuded latex was studied using cytochemistry. Articulated laticifers were present in M. atroviolacea. The lactiferous system in stems and leaves originated from both procambium and ground meristem, and it was determined to be functional, as indicated by changes in the cytoplasm immediately subsequent to its formation. However, in tuberous roots, laticifers originated from the vascular cambium, phellogen, and parenchymatous cortical cells. The primary wall of laticifers consists of pectins and cellulose and is thicker than that of adjacent cells. The laticifer secretion contained an emulsion of lipophilic substances rich in neutral lipids, resin oils, and possibly rubber. The presence of articulated laticifers is reported for the first time for Apocynoideae, contributing to the knowledge and understanding of these structures in the family Apocynaceae. Key words: Apocynaceae, Mandevilla atroviolacea, articulated laticifers, latex, ontogeny. Dans la litterature, on a enregistre la presence de laticiferes non articules chez les Apocynaceae, mais on trouve peu d'information sur la structure et la composition chimiques des composes du latex. L'objectif de cette etude consistait a decrire l'anatomie et l'histochimie des laticiferes et du latex des racines, des tiges et des feuilles du Mandevilla atroviolacea (Stadelm) Woodson, une espece d' Apocynaceae. Les auteurs ont examine l'anatomie des laticiferes a l'aide de la microscopie photonique et la nature chimique du latex exsude par cytochimie. On retrouve des laticiferes articules chez le M. atroviolacea. Le systeme laticifere des tiges et des feuilles provient a la fois du procambium et du meristeme fondamental, et des changements dans le cytoplasme, immediatement apres sa formation, indiquent sa fonctionnalite. Cependant, dans les racines tubereuses, les laticiferes proviennent du cambium vasculaire, du phellogene et des cellules du parenchyme cortical. La paroi primaire des laticiferes, plus epaisse que celle des cellules adjacentes, est constituee de pectine et de cellulose. La secretion des laticiferes contient une emulsion de substances lipophiles riches en lipides neutres, en huiles de resine et possiblement en caoutchouc. Les auteurs rapportent pour la premiere fois la presence de laticiferes articules chez les Apocynoideae, contribuant ainsi a la connaissance et a la comprehension de ces structures chez les Apocynaceae. Mots-cles : Apocynaceae, Mandevillea atroviolacea, laticiferes articules, latex, ontogenie. [Traduit par la Redaction], Introduction Laticifers are found in approximately 900 genera and 20 families, mostly dicotyledonous species but also in a few monocotyledonous species (Solereder 1908; Metcalfe 1967; Fahn 1979). Therefore, they are [...]

In stoloniferous species, the length of petioles is of pivotal importance because it determines the position of leaf blades within the canopy. From a mechanistic perspective, two developmental processes, cell division and cell elongation, are responsible for the length of a given petiole. This study aimed at quantifying the relative contributions of cell division and cell elongation to genotypic and plastic variation in petiole length of the stoloniferous herb Trifolium repens. Thirty-four genotypes of T. repens were grown under high light conditions and simulated canopy shade. Cells were counted and their lengths measured on epidermal prints from fully grown petioles of leaves that had been initiated in the experimental light conditions. Cell number was the main trait explaining petiole length differences among genotypes grown under high light, while both cell number and length changed in response to shading. Our study revealed a strong negative correlation between shade-induced changes in cell number and cell length: genotypes that responded to shading by increasing cell numbers hardly changed in cell length, and vice versa. Our results suggest that genotypic and phenotypic variation in petiole length results from a complex interplay between the developmental processes of cell elongation and cell division. Key words: cell number; cell size; petiole elongation; plasticity; shade avoidance; trade-off; Trifolium repens.

Production of high-value recombinant proteins in transgenic seeds is an attractive and economically feasible alternative to conventional systems based on mammalian cells and bacteria. In contrast to leaves, seeds allow high-level accumulation of recombinant proteins in a relatively small volume and a stable environment. We demonstrate that single-chain variable fragment (scFv)-Fc antibodies, with N-terminal signal sequence and C-terminal KDEL tag, can accumulate to very high levels as bivalent IgG-like antibodies in Arabidopsis thaliana seeds and illustrate that a plant-produced anti-hepatitis A virus scFv-Fc has similar antigen-binding and in vitro neutralizing activities as the corresponding full-length IgG. As expected, most scFv-Fc produced in seeds contained only oligomannose-type N-glycans, but, unexpectedly, 35-40% was never glycosylated. A portion of the scFv-Fc was found in endoplasmic reticulum (ER)-derived compartments delimited by ribosome-associated membranes. Additionally, consistent with the glycosylation data, large amounts of the recombinant protein were deposited in the periplasmic space, implying a direct transport from the ER to the periplasmic space between the plasma membrane and the cell wall. Aberrant localization of the ER chaperones calreticulin and binding protein (BiP) and the endogenous seed storage protein cruciferin in the periplasmic space suggests that overproduction of recombinant scFv-Fc disturbs normal ER retention and protein-sorting mechanisms in the secretory pathway. glycosylation | molecular farming | recombinant antibody | subcellular localization

Antioxidant enzymes protect plant cells from oxidative injury induced by hypoxia. The objective of this study was to identify the responses of antioxidant enzymes to different depths of waterlogging (WL) in creeping bentgrass (Agrostis stolonifera L.) roots. 'Penncross' and 'G-6' were subjected to 21 d of WL at 1 cm (WL-1) and 15 cm (WL-15) below the soil surface, respectively. The turf quality of Penncross was not acceptable under both WL conditions, while G-6 had an acceptable quality of 6.5 under WL-15. The activity of superoxide dismutase (SOD) increased by 32 and 26% for Penncross and 83 and 44% for G-6 under WL-15 and WL-1, respectively. One isoform of Mn-SOD and three isoforms of Cu/Zn-SOD were identified in both cultivars under all treatments. The activity of ascorbate peroxidase (AP) significantly decreased by 46 and 40% for Penncross, while AP activity decreased 25% and increased 9% for G-6 under WL-1 and WL-15, respectively. One isoform of AP was identified with strong intensity in both cultivars under all treatments. The content of hydrogen peroxide ([H.sub.2][O.sub.2]) and malondialdehyde (MDA, 1,1,3,3-tetramethoxy-propane) and the activities of glutathione reductase (GR) and peroxidase (POD) were not affected by WL for both cultivars. These results indicated that SOD and AP were mainly involved in WL-induced antioxidant responses, and the partial WL could also significantly affect root antioxidant activities, particularly in WL-sensitive cultivar.

From a mechanical point of view, plant and hyphal cells are more complex than their animal counterparts because the variety of structural components determining cellular architecture is broader. In addition to cytoskeletal elements and the plasma membrane, the cell wall and turgor pressure equip plant and hyphal cells with structures analogous to an exoskeleton and a hydroskeleton, respectively. To quantify the physical properties of plant and hyphal cells, researchers have developed a plethora of experimental methods. This review provides an overview of experimental approaches that have been used to measure turgor pressure and to determine the mechanical properties of the plant cell wall at the subcellular level. It is completed by a glimpse into the arsenal of techniques that has been used to characterize the physical properties of cytoskeletal elements. These have mostly been used on animal cells, but we hope they will find their way into plant cell research. Finally, assays and tests to measure the generation of forces by cells and subcellular structures are discussed. Key words: cell biomechanics; cell wall; cytomechanics; cytoskeleton; hypha; plant cell; tip growth; turgor.

The bacterial plant pathogen Pseudomonas syringae requires a type III protein secretion system (TTSS) to cause disease. The P. syringae TTSS is encoded by the hrp-hrc gene cluster. One of the genes within this cluster, hrpJ, encodes a protein with weak similarity to YopN, a type III secreted protein from the animal pathogenic Yersinia species. Here, we show that HrpJ is secreted in culture and translocated into plant cells by the P. syringae pv. tomato DC3000 TTSS. A DC3000 hrpJ mutant, UNL140, was greatly reduced in its ability to cause disease symptoms and multiply in Arabidopsis thaliana. UNL140 exhibited a reduced ability to elicit a hypersensitive response (HR) in nonhost tobacco plants. UNL140 was unable to elicit an AvrRpt2- or AvrB1-dependent HR in A. thaliana but maintained its ability to secrete AvrB1 in culture via the TTSS. Additionally, UNL140 was defective in its ability to translocate the effectors AvrPtol, HopB1, and AvrPtoB. Type III secretion assays showed that UNL140 secreted HrpA1 and AvrPtol but was unable to secrete HrpZ1, a protein that is normally secreted in culture in relatively large amounts, into culture supernatants. Taken together, our data indicate that HrpJ is a type III secreted protein that is important for pathogenicity and the translocation of effectors into plant cells. Based on the failure of UNL140 to secrete HrpZ1, HrpJ may play a role in controlling type III secretion, and in its absence, specific accessory proteins, like HrpZ1, may not be extracellularly localized, resulting in disabled translocation of effectors into plant cells.

We propose a theoretical mechanism that enables the elaboration of veins to supply distant cells during leaf development. In contrast to the more standard view that a signal (e.g., auxin) is produced at isolated sites to stimulate growth, we determine the consequences of the hypothesis that auxin is produced at a constant rate in every cell. High concentration sites for auxin emerge naturally in a reaction-diffusion model, together with global information about leaf shape and existing venation. Because the global information is encoded as auxin concentration and its gradient, those signals provide individual cells with sufficient information to determine their own fate. Unlike other models, a single substance suffices for the reaction-diffusion at early, but not initial, stages of development. Neither complex interactions nor predetermination are necessary. We predict angiosperm areolation patterns in simulation, and our model further implies the Sachs Canalization Hypothesis and resolves a dilemma regarding the role of auxin in cell growth. canalization hypothesis | emergent behavior | reaction-diffusion