The AGCVIII kinase Dw2 modulates cell proliferation, endomembrane trafficking, and MLG/xylan cell wall localization in elongating stem internodes of Sorghum bicolor.

Stems of bioenergy sorghum (Sorghum bicolor L. Moench.), a drought-tolerant C4 grass, contain up to 50 nodes and internodes of varying length that span 4-5 m and account for approximately 84% of harvested biomass. Stem internode growth impacts plant height and biomass accumulation and is regulated by brassinosteroid signaling, auxin transport, and gibberellin biosynthesis. In addition, an AGCVIII kinase (Dw2) regulates sorghum stem internode growth, but the underlying mechanism and signaling network are unknown. Here we provide evidence that mutation of Dw2 reduces cell proliferation in internode intercalary meristems, inhibits endocytosis, and alters the distribution of heteroxylan and mixed linkage glucan in cell walls. Phosphoproteomic analysis showed that Dw2 signaling influences the phosphorylation of proteins involved in lipid signaling (PLDδ), endomembrane trafficking, hormone, light, and receptor signaling, and photosynthesis. Together, our results show that Dw2 modulates endomembrane function and cell division during sorghum internode growth, providing insight into the regulation of monocot stem development.

The Metabolite Pathway between Bundle Sheath and Mesophyll: Quantification of Plasmodesmata in Leaves of C3 and C4 Monocots.

C4 photosynthesis is characterized by a CO2-concentrating mechanism between mesophyll (M) and bundle sheath (BS) cells of leaves. This generates high metabolic fluxes between these cells, through interconnecting plasmodesmata (PD). Quantification of these symplastic fluxes for modeling studies requires accurate quantification of PD, which has proven difficult using transmission electron microscopy. Our new quantitative technique combines scanning electron microscopy and 3D immunolocalization in intact leaf tissues to compare PD density on cell interfaces in leaves of C3 (rice [Oryza sativa] and wheat [Triticum aestivum]) and C4 (maize [Zea mays] and Setaria viridis) monocot species. Scanning electron microscopy quantification of PD density revealed that C4 species had approximately twice the number of PD per pitfield area compared with their C3 counterparts. 3D immunolocalization of callose at pitfields using confocal microscopy showed that pitfield area per M-BS interface area was 5 times greater in C4 species. Thus, the two C4 species had up to nine times more PD per M-BS interface area (S. viridis, 9.3 PD µm(-2); maize, 7.5 PD µm(-2); rice 1.0 PD µm(-2); wheat, 2.6 PD µm(-2)). Using these anatomical data and measured photosynthetic rates in these C4 species, we have now calculated symplastic C4 acid flux per PD across the M-BS interface. These quantitative data are essential for modeling studies and gene discovery strategies needed to introduce aspects of C4 photosynthesis to C3 crops.

Out-of-plane orientation of cellulose elementary fibrils on spruce tracheid wall based on imaging with high-resolution transmission electron microscopy.

MAIN CONCLUSION: A 3D model of the tracheid wall has been proposed based on high-resolution cryo-TEM where, in contrast to the current understanding, the cellulose elementary fibrils protrude from the cell wall plane. The ultrastructure of the tracheid walls of Picea abies was examined through imaging of ultrathin radial, tangential and transverse sections of wood by transmission electron microscopy and with digital image processing. It was found that the elementary fibrils (EFs) of cellulose were rarely deposited in the plane of the concentric cell wall layers, in contrast to the current understanding. In addition to the adopted concept of longitudinal fibril angle, EFs protruded from the cell wall plane in varying angles depending on the layer. Moreover, the out-of-plane fibril angle varied between radial and tangential walls. In the tangential S2 layers, EFs were always out-of-plane whereas planar orientation was typical for the S2 layer in radial walls. The pattern of protruding EFs was evident in almost all axial and transverse images of the S1 layer. Similar out-of-plane orientation was found in the transverse sections of the S3 layer. A new model of the tracheid wall with EF orientation is presented as a summary of this study. The outcome of this study will enhance our understanding of the elementary fibril orientation in the tracheid wall.

Vascular occlusions in grapevines with Pierce's disease make disease symptom development worse.

Vascular occlusions are common structural modifications made by many plant species in response to pathogen infection. However, the functional role(s) of occlusions in host plant disease resistance/susceptibility remains controversial. This study focuses on vascular occlusions that form in stem secondary xylem of grapevines (Vitis vinifera) infected with Pierce's disease (PD) and the impact of occlusions on the hosts' water transport and the systemic spread of the causal bacterium Xylella fastidiosa in infected vines. Tyloses are the predominant type of occlusion that forms in grapevine genotypes with differing PD resistances. Tyloses form throughout PD-susceptible grapevines with over 60% of the vessels in transverse sections of all examined internodes becoming fully blocked. By contrast, tylose development was mainly limited to a few internodes close to the point of inoculation in PD-resistant grapevines, impacting only 20% or less of the vessels. The extensive vessel blockage in PD-susceptible grapevines was correlated to a greater than 90% decrease in stem hydraulic conductivity, compared with an approximately 30% reduction in the stems of PD-resistant vines. Despite the systemic spread of X. fastidiosa in PD-susceptible grapevines, the pathogen colonized only 15% or less of the vessels in any internode and occurred in relatively small numbers, amounts much too small to directly block the vessels. Therefore, we concluded that the extensive formation of vascular occlusions in PD-susceptible grapevines does not prevent the pathogen's systemic spread in them, but may significantly suppress the vines' water conduction, contributing to PD symptom development and the vines' eventual death.

Initial events during the evolution of C4 photosynthesis in C3 species of Flaveria.

The evolution of C4 photosynthesis in many taxa involves the establishment of a two-celled photorespiratory CO2 pump, termed C2 photosynthesis. How C3 species evolved C2 metabolism is critical to understanding the initial phases of C4 plant evolution. To evaluate early events in C4 evolution, we compared leaf anatomy, ultrastructure, and gas-exchange responses of closely related C3 and C2 species of Flaveria, a model genus for C4 evolution. We hypothesized that Flaveria pringlei and Flaveria robusta, two C3 species that are most closely related to the C2 Flaveria species, would show rudimentary characteristics of C2 physiology. Compared with less-related C3 species, bundle sheath (BS) cells of F. pringlei and F. robusta had more mitochondria and chloroplasts, larger mitochondria, and proportionally more of these organelles located along the inner cell periphery. These patterns were similar, although generally less in magnitude, than those observed in the C2 species Flaveria angustifolia and Flaveria sonorensis. In F. pringlei and F. robusta, the CO2 compensation point of photosynthesis was slightly lower than in the less-related C3 species, indicating an increase in photosynthetic efficiency. This could occur because of enhanced refixation of photorespired CO2 by the centripetally positioned organelles in the BS cells. If the phylogenetic positions of F. pringlei and F. robusta reflect ancestral states, these results support a hypothesis that increased numbers of centripetally located organelles initiated a metabolic scavenging of photorespired CO2 within the BS. This could have facilitated the formation of a glycine shuttle between mesophyll and BS cells that characterizes C2 photosynthesis.

Was low CO2 a driving force of C4 evolution: Arabidopsis responses to long-term low CO2 stress.

The responses of long-term growth of plants under elevated CO2 have been studied extensively. Comparatively, the responses of plants to subambient CO2 concentrations have not been well studied. This study aims to investigate the responses of the model C3 plant, Arabidopsis thaliana, to low CO2 at the molecular level. Results showed that low CO2 dramatically decreased biomass productivity, together with delayed flowering and increased stomatal density. Furthermore, alteration of thylakoid stacking in both bundle sheath and mesophyll cells, upregulation of PEPC and PEPC-K together with altered expression of a number of regulators known involved in photosynthesis development were observed. These responses to low CO2 are discussed with regard to the fitness of C3 plants under low CO2. This work also briefly discusses the relevance of the data to C4 photosynthesis evolution.

Bundle sheath suberization in grass leaves: multiple barriers to characterization.

High-yielding, stress-tolerant grass crops are essential to meet future food and energy demands. Efforts are underway to engineer improved varieties of the C3 cereal crop rice by introducing NADP-malic enzyme C4 photosynthesis using maize as a model system. However, several modifications to the rice leaf vasculature are potentially necessary, including the introduction of suberin lamellae into the bundle sheath cell walls. Suberized cell walls are ubiquitous in the root endodermis of all grasses, and developmental similarities are apparent between endodermis and bundle sheath cell walls. Nonetheless, there is considerable heterogeneity in sheath cell development and suberin composition both within and between grass taxa. The effect of this variation on physiological function remains ambiguous over forty years after suberin lamellae were initially proposed to regulate solute and photoassimilate fluxes and C4 gas exchange. Interspecies variation has confounded efforts to ascribe physiological differences specifically to the presence or absence of suberin lamellae. Thus, specific perturbation of suberization within a uniform genetic background is needed, but, until recently, the genetic resources to manipulate suberin composition in the grasses were largely unavailable. The recent dissection of the suberin biosynthesis pathway in model dicots and the identification of several promising candidate genes in model grasses will facilitate the characterization of the first suberin biosynthesis genes in a monocot. Much remains to be learned about the role of bundle sheath suberization in leaf physiology, but the stage is set for significant advances in the near future.

[Structure, ontogeny and vascularization of the flowers and inflorescences of Drimys granadensis (Winteraceae)]. / Estructura, ontogenia y vascularización de las flores e inflorescencias de Drimys granadensis (Winteraceae).

Structure, ontogeny and vascularization of the flowers and inflorescences of Drimys granadensis (Winteraceae). Drimys granadensis is a widespread species in montane forests of South and Central America. In this research, the structure, ontogeny, phyllotaxis and vascularization of the flowers and inflorescences of this species was studied in a population from the Eastern hills of Sabana de Bogota, Colombia. The methods used applied both optical microscopy, with astra blue-fuchsin staining, and scanning electron microscopy, using critical point dryed and gold-paladium metallized samples. Besides, results were compared with those of Drimys winteri, a widely studied species distributed in Chile and Argentina. Additionally, we studied the detail of the floral anatomy to determine the bracteal or calicine identity of the caliptra. I confirmed the proliferative status of the monothelic inflorescence, discarding alternative explanations of the terminal flower identity. I found that uniflorescences have an acropetal development until the terminal meristem becomes the terminal flower, then this flower develops rapidly resulting in a determined uniflorescence. I found pseudosyphonosthelic vascularization in peduncles and pedicels. Besides, I discovered some evidence in the vascular and anatomical structures, to consider the caliptra as the fusion product of various structures and therefore of calicine origin. The caliptra showed a whorled phyllotaxis, but the petals, stamens and carpels presented a spiral condition; phyllotaxis change was explained by the long time lapse between the initiation of the calyx and the corolla. I found great similarities among the inflorescences of D. granadensis and D. winteri; they were different in the proliferation start time, and in the frequent presence of nomophylls in D. granadensis, in contrast to the presence of reduced bracts and bracteoles in D. winteri inflorescences.

Individual maize chromosomes in the C(3) plant oat can increase bundle sheath cell size and vein density.

C(4) photosynthesis has evolved in at least 66 lineages within the angiosperms and involves alterations to the biochemistry, cell biology, and development of leaves. The characteristic "Kranz" anatomy of most C(4) leaves was discovered in the 1890s, but the genetic basis of these traits remains poorly defined. Oat × maize addition lines allow the effects of individual maize (Zea mays; C(4)) chromosomes to be investigated in an oat (Avena sativa; C(3)) genetic background. Here, we have determined the extent to which maize chromosomes can introduce C(4) characteristics into oat and have associated any C(4)-like changes with specific maize chromosomes. While there is no indication of a simultaneous change to C(4) biochemistry, leaf anatomy, and ultrastructure in any of the oat × maize addition lines, the C(3) oat leaf can be modified at multiple levels. Maize genes encoding phosphoenolpyruvate carboxylase, pyruvate, orthophosphate dikinase, and the 2'-oxoglutarate/malate transporter are expressed in oat and generate transcripts of the correct size. Three maize chromosomes independently cause increases in vein density, and maize chromosome 3 results in larger bundle sheath cells with increased cell wall lipid deposition in oat leaves. These data provide proof of principle that aspects of C(4) biology could be integrated into leaves of C(3) crops.

Antibodies against Yariv's reagent for immunolocalization of arabinogalactan-proteins in aerial parts of Echinacea purpurea.

Arabinogalactan-proteins are glycoproteins that occur in higher plants and are involved in important processes like cell differentiation and plant growth. In the medicinal plant Echinacea purpurea L., they belong to the putative immunomodulating compounds and are structurally well characterized. For microscopic localization of arabinogalactan-proteins, synthetic (ß-D-Glc)3 Yariv phenylglycoside that specifically binds to most plant arabinogalactan-proteins was used to label arabinogalactan-proteins in fresh cut sections of stems and petioles of Echinacea purpurea. Polyclonal antibodies against (ß-D-Glc)3 Yariv phenylglycoside were used to detect the arabinogalactan-protein-(ß-D-Glc)3 Yariv phenylglycoside complex. After addition of fluorescein isothiocyanate-conjugated secondary antibodies, the sections were analyzed by confocal laser scanning microscopy. Arabinogalactan-proteins are localized mainly in the central cylinder in the collateral vascular bundles, especially in the area of the xylem. In cell walls of fully differentiated vessels and tracheids, arabinogalactan-proteins have been detected mainly at the inner area of the wall close to the cell lumina. Intense labeling occurs around pit canals connecting adjacent vessels. Furthermore, arabinogalactan-proteins are present in the lumina of cells of the sclerenchyma caps and in companion cells of the phloem.

Immunolocalization of hemicelluloses in Arabidopsis thaliana stem. Part I: temporal and spatial distribution of xylans.

We investigated the microdistribution of xylans in different cell types of Arabidopsis stem using immunolocalization methods with LM10 and LM11 antibodies. Xylan labeling in xylary fibers (fibers) was initially detected at the cell corner of the S(1) layer and increased gradually during fiber maturation, showing correlation between xylan labeling and general secondary cell wall formation processes in fibers. Metaxylem vessels (vessels) showed earlier development of secondary cell walls than fibers, but revealed almost identical labeling patterns to fibers during maturation. No difference in labeling patterns and intensity was detected in the cell wall of fibers, vessels and protoxylem vessels (proto-vessels) between LM10 and LM11, indicating that vascular bundle cells may be chemically composed of a highly homogeneous xylan type. Interestingly, interfascicular fibers (If-fibers) showed different labeling patterns between the two antibodies and also between different developmental stages. LM10 showed no labeling in primary cell walls and intercellular layers of If-fibers at the S(1) formation stage, but some labeling was detected in middle lamella cell corner regions at the S(2) formation stage. In contrast, LM11 revealed uniform labeling across the If-fiber cell wall during all developmental stages. These results suggest that If-fibers have different xylan deposition processes and patterns from vascular bundle cells. The presence of xylan was also confirmed in parenchyma cells following pectinase treatment. Together our results indicate that there are temporal and spatial differences in xylan labeling between cell types in Arabidopsis stem. Differences in xylan labeling between Arabidopsis stem and poplar are also discussed.

Overexpression of phytochrome A and its hyperactive mutant improves shade tolerance and turf quality in creeping bentgrass and zoysiagrass.

Phytochrome A (phyA) in higher plants is known to function as a far-red/shade light-sensing photoreceptor in suppressing shade avoidance responses (SARs) to shade stress. In this paper, the Avena PHYA gene was introduced into creeping bentgrass (Agrostis stolonifera L.) and zoysiagrass (Zoysia japonica Steud.) to improve turf quality by suppressing the SARs. In addition to wild-type PHYA, a hyperactive mutant gene (S599A-PHYA), in which a phosphorylation site involved in light-signal attenuation was removed, was also transformed into the turfgrasses. Phenotypic traits of the transgenic plants were compared to assess the suppression of SARs under a simulated shade condition and outdoor field conditions after three growth seasons. Under the shade condition, the S599A-PhyA transgenic creeping bentgrass plants showed shade avoidance-suppressing phenotypes with a 45 % shorter leaf lengths, 24 % shorter internode lengths, and twofold increases in chlorophyll concentrations when compared with control plants. Transgenic zoysiagrass plants overexpressing S599A-PHYA also showed shade-tolerant phenotypes under the shade condition with reductions in leaf length (15 %), internode length (30 %), leaf length/width ratio (19 %) and leaf area (22 %), as well as increases in chlorophyll contents (19 %) and runner lengths (30 %) compared to control plants. The phenotypes of transgenic zoysiagrass were also investigated in dense field habitats, and the transgenic turfgrass exhibited shade-tolerant phenotypes similar to those observed under laboratory shade conditions. Therefore, the present study suggests that the hyperactive phyA is effective for the development of shade-tolerant plants, and that the shade tolerance nature is sustained under field conditions.

Molecular cloning and characterization of OsCHR4, a rice chromatin-remodeling factor required for early chloroplast development in adaxial mesophyll.

Mi-2 protein, the central component of the NuRD nucleosome remodeling and histone deacetylase complex, plays a role in transcriptional repression in animals. Mi-2-like genes have been reported in Arabidopsis, though their function in monocots remains largely unknown. In the present study, a rice Mi-2-like gene, OsCHR4 (Oryza sativa Chromatin Remodeling 4, LOC_Os07g03450), was cloned from a rice mutant with adaxial albino leaves. The Oschr4 mutant exhibited defective chloroplasts in adaxial mesophyll, but not in abaxial mesophyll. Ultrastructural observations indicated that proplastid growth and/or thylakoid membrane formation in adaxial mesophyll cells was blocked in the Oschr4 mutant. Subcellular localization revealed that OsCHR4::GFP fusion protein was targeted to the nuclei. OsCHR4 was mainly expressed in the root meristem, flower, vascular bundle, and mesophyll cells by promoter::GUS analysis in transgenic rice. The transcripts of some nuclear- and plastid-encoded genes required for early chloroplast development and photosynthesis were decreased in the adaxial albino mesophyll of the Oschr4 mutant. These observations provide evidence that OsCHR4, the rice Mi-2-like protein, plays an important role in early chloroplast development in adaxial mesophyll cells. The results increase our understanding of the molecular mechanism underlying tissue-specific chloroplast development in plants.

Imaging cell wall architecture in single Zinnia elegans tracheary elements.

The chemical and structural organization of the plant cell wall was examined in Zinnia elegans tracheary elements (TEs), which specialize by developing prominent secondary wall thickenings underlying the primary wall during xylogenesis in vitro. Three imaging platforms were used in conjunction with chemical extraction of wall components to investigate the composition and structure of single Zinnia TEs. Using fluorescence microscopy with a green fluorescent protein-tagged Clostridium thermocellum family 3 carbohydrate-binding module specific for crystalline cellulose, we found that cellulose accessibility and binding in TEs increased significantly following an acidified chlorite treatment. Examination of chemical composition by synchrotron radiation-based Fourier-transform infrared spectromicroscopy indicated a loss of lignin and a modest loss of other polysaccharides in treated TEs. Atomic force microscopy was used to extensively characterize the topography of cell wall surfaces in TEs, revealing an outer granular matrix covering the underlying meshwork of cellulose fibrils. The internal organization of TEs was determined using secondary wall fragments generated by sonication. Atomic force microscopy revealed that the resulting rings, spirals, and reticulate structures were composed of fibrils arranged in parallel. Based on these combined results, we generated an architectural model of Zinnia TEs composed of three layers: an outermost granular layer, a middle primary wall composed of a meshwork of cellulose fibrils, and inner secondary wall thickenings containing parallel cellulose fibrils. In addition to insights in plant biology, studies using Zinnia TEs could prove especially productive in assessing cell wall responses to enzymatic and microbial degradation, thus aiding current efforts in lignocellulosic biofuel production.

Bundle sheath suberisation is required for C4 photosynthesis in a Setaria viridis mutant.

C4 photosynthesis provides an effective solution for overcoming the catalytic inefficiency of Rubisco. The pathway is characterised by a biochemical CO2 concentrating mechanism that operates across mesophyll and bundle sheath (BS) cells and relies on a gas tight BS compartment. A screen of a mutant population of Setaria viridis, an NADP-malic enzyme type C4 monocot, generated using N-nitroso-N-methylurea identified a mutant with an amino acid change in the gene coding region of the ABCG transporter, a step in the suberin synthesis pathway. Here, Nile red staining, TEM, and GC/MS confirmed the alteration in suberin deposition in the BS cell wall of the mutant. We show that this has disrupted the suberin lamellae of BS cell wall and increased BS conductance to CO2 diffusion more than two-fold in the mutant. Consequently, BS CO2 partial pressure is reduced and CO2 assimilation was impaired in the mutant. Our findings provide experimental evidence that a functional suberin lamellae is an essential anatomical feature for efficient C4 photosynthesis in NADP-ME plants like S. viridis and have implications for engineering strategies to ensure future food security.

Extraction and characterization of vascular bundle and fiber strand from date palm rachis as potential bio-reinforcement in composite.

Date palm rachis fibers are rich in cellulose, relatively inexpensive, and readily available in Algeria. The aim of this study is to investigate the morphology, structure, mechanical and physicochemical characteristics of both vascular bundles and fiber strands extracted from date palm rachis. The difficulties encountered are associated to the extraction of the fibers without damaging them. The study focuses on the morphological and surface roughness analysis using optical and scanning electron microscopies (SEM), and a non-contact 3D profiler. The chemical, physical and thermal properties have been studied using Fourier-transform infrared (FTIR) spectroscopy, energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). The mechanical properties were accessed by tensile tests and they were analyzed using two-parameter Weibull distribution.

Floral nectaries in Sapindaceae s.s.: morphological and structural diversity, and their systematic implications.

We investigated the morphology and structure of the floral nectary in 11 Neotropical genera belonging to the subfamilies Dodonaeoideae and Paullinioideae (Sapindaceae) from southern South America representing three tribes (Dodonaeaeae, Paullinieae, and Melicocceae), in relation to other floral traits in species with contrasting morphological flower characteristics. Nectary organization was analyzed under light, stereoscopic, and scanning electron microscopes; Diplokeleba floribunda N.E. Br. was also observed using transmission electron microscopy. Our comparative data may contribute to the understanding of floral nectary evolution and systematic value in this family. The nectaries were studied in both staminate and pistillate flowers. All the floral nectaries are typical of Sapindaceae: extrastaminal, receptacular, structured, and persistent. The anatomical analysis revealed a differentiated secretory parenchyma and an inner non-secretory parenchyma; the nectary is supplied by phloem traces and, less frequently, by phloem and xylem traces. Nectar is secreted through nectarostomata of anomocytic type. The anatomical analysis showed the absence of nectary in the three morphs of Dodonaea viscosa flowers. Nectary ultrastructure is described in D. floribunda. In this species, the change in nectary color is related to progressive accumulation of anthocyanins during the functional phase. We found relatively small variation in the nectary structural characteristics compared with large variation in nectary morphology. The latter aspect agreed with the main infrafamilial groupings revealed by recent phylogenetic studies, so it is of current valuable systematic importance for Sapindaceae. In representatives of Paullinieae, the reduction of the floral nectary to 4-2 posterior lobes should be interpreted as a derived character state.

Cold induced changes in the water balance affect immunocytolocalization pattern of one of the aquaporins in the vascular system in the leaves of maize (Zea mays L.).

Chilling stress is known to affect the water balance in plants, which often manifests itself in the decrease of the water potential in different organs. Relationships between chilling, assimilate transport and water balance are far from being understood. Although aquaporins play a key role in regulating water balance in plants, especially under stress conditions, the role of individual aquaporins in stress response remains unclear. In this report we show the specific localization within plasma membranes of one of the aquaporins (PIP2;3) in the leaves of two maize inbred lines differing in their chilling-sensitivity. This form of aquaporin has been also observed in thick-walled sieve elements - an additional type of sieve tubes of unclear function found only in monocotyledons. Moderate chilling (about 15°C) caused significant reduction of labelling in these cells accompanied by a steep decrease in the water potential in leaves of chilling-sensitive maize line. Our results suggest that both PIP2;3 and thick-walled sieve tubes may be an unknown element of the mechanism of the response of maize to cold stress.

Observation and investigation of three endosperm transport tissues in sorghum caryopses.

Endosperm transport tissues in sorghum caryopses include endosperm transfer cells, endosperm conducting cells, and the embryo surrounding region. To elucidate the structural changes of these tissues and their relationship with the caryopsis development, sorghum caryopses were analyzed at different days after pollination using light, fluorescence, and electron microscopy. The following results were obtained: post-phloem maternal tissues included the placentochalaza and the nucellar projection-like nucellus. Well-developed endosperm transfer cells exhibited very evident flange-type wall ingrowths. Very few wall ingrowths were present in the initially developed endosperm transfer cells when the level of sucrose from the initially developed vascular system was low. At the middle stage of caryopsis development, the level of sucrose from the well-developed vascular system was high. Endosperm transfer cells increased in both area and layer amount, and their wall ingrowths increased in both length and density. Later in caryopsis development, the level of sucrose from the degenerated vascular system was low and wall ingrowths distorted in the degenerated endosperm transfer cells. Endosperm conducting cells primarily occupied the most part of endosperm, but decreased gradually because the upper part transformed into the starchy endosperm and the lower part degenerated to give space to the embryo growth. Although the embryo surrounding region initially enveloped the small embryo, it rapidly degenerated and finally disappeared. Our data showed that (1) the caryopsis vascular system influenced the differentiation of endosperm transfer cells by controlling the sugar levels (2) and configuration of endosperm transport tissues were probably altered to favor the growth of filial tissues.

Disruption of OsEXO70A1 Causes Irregular Vascular Bundles and Perturbs Mineral Nutrient Assimilation in Rice.

Normal uptake, transportation, and assimilation of primary nutrients are essential to plant growth. Tracheary elements (TEs) are tissues responsible for the transport of water and minerals and characterized by patterned secondary cell wall (SCW) thickening. Exocysts are involved in the regulation of SCW deposition by mediating the targeted transport of materials and enzymes to specific membrane areas. EXO70s are highly duplicated in plants and provide exocysts with functional specificity. In this study, we report the isolation of a rice mutant rapid leaf senescence2 (rls2) that exhibits dwarfism, ferruginous spotted necrotic leaves, decreased hydraulic transport, and disordered primary nutrient assimilation. Histological analysis of rls2-1 mutants has indicated impaired cell expansion, collapsed vascular tissues, and irregular SCW deposition. Map-based cloning has revealed that RLS2 encodes OsEXO70A1, which is one of the 47 members of EXO70s in rice. RLS2 was widely expressed and spatially restricted in vascular bundles. Subcellular localization analysis demonstrated that RLS2 was present on both membrane and nuclear regions. Expression analysis revealed that mutations in rls2 triggers transcriptional fluctuation of orthologous EXO70 genes and affects genes involved in primary nutrient absorption and transport. In brief, our study revealed that RLS2 is required for normal vascular bundle differentiation and primary nutrient assimilation.