Plasma Membrane Of Epidermal Cells Research Articles

ABCG11 modulates cytokinin responses in Arabidopsis thaliana.

The Arabidopsis ABC transporter ABCG11 transports lipidic precursors of surface coating polymers at the plasma membrane of epidermal cells. Mutants in ABCG11 exhibit severe developmental defects, suggesting that ABCG11 might also participate in phytohormone-mediated development. Here, we report that ABCG11 is involved in cytokinin-mediated development. The roots of abcg11 mutant seedlings failed to respond to cytokinins and accumulated more cytokinins than wild-type roots. When grown under short-day conditions, abcg11 exhibited longer roots and shorter hypocotyls compared to wild type, similar to abcg14, a knockout mutant in a cytokinin transporter. Treatment with exogenous trans-zeatin, which inhibits primary root elongation in the wild type, enhanced abcg11 primary root elongation. It also increased the expression of cytokinin-responsive Arabidopsis response regulator (ARR) genes, and the signal of the TCS::GFP reporter in abcg11 roots compared to wild-type roots, suggesting that cytokinin signaling was enhanced in abcg11 roots. When we treated only the roots of abcg11 with trans-zeatin, their shoots showed lower ARR induction than the wild type. The abcg14 abcg11 double mutant did not have additional root phenotypes compared to abcg11. Together, these results suggest that ABCG11 is necessary for normal cytokinin-mediated root development, likely because it contributes to cytokinin transport, either directly or indirectly.

UVB Dose Optimization for Phototherapy in Vitamin D Deficiency : Profile Analysis of Vitamin D, TNF-α, Vascular Endothelial Growth Factor (VEGF) and Platelet Derived Growth Factor (PDGF) in Wistar Rats

Background : UVB radiation responsible for the most important biological effects including Vitamin D3 synthesis and inflammation. UVB radiation are absorbed by 7-dehydrocholesterol in the plasma membrane of epidermal cells resulting in production of cis-previtamin D3. In the other hand, an exposure to UVB leads to cutaneous tissue inflammation modulates by TNF-α which also increases platelet activating factor. VEGF and PDGF induced by TNF-α during wound healing, characterized with angiogenesis and reephitalization. Furthermore, vitamin D plays a role in inflammation inhibition and upregulates growth factors. However, the study of the mechanism has not yet been thoroughly investigated.
 Methods: This study uses post test only group design, subjected wistar rats divided into four groups. Control group, non irradiated with UVB, and the other three groups, treated with graded UVB dose started with 1 MED (50 mJ/cm2), 2 MED (100mJ/cm2) and 3 MED (150 mJ/cm2) and investigated at 6, 12, 24 and 48 hours post UVB irradiation.
 Result : The serum level of vitamin D, VEGF and PDGF were increasing due to UVB dose addition. The highest level was reached at 6 hours post radiation using 3 MED, which gradually decrease up to 48 hours (p =0,000). The rise of vitamin D after UVB radiation, inhibit TNF-α induction in every dose accordant UVB dose addition and the lowest level is using 3 MED at 12 hours post radiation (p =0,000). TNF-α reach its highest level at 24 hours post radiation using 1 MED, it is related with the acute phase of inflammation.
 Conclusion : This study reveal that higher UVB irradiance increases vitamin D and inhibit TNF-α which also promotes VEGF and PDGF.
 Bangladesh Journal of Medical Science Vol.19(4) 2020 p.749-754

MsPG1 alleviated aluminum-induced inhibition of root growth by decreasing aluminum accumulation and increasing porosity and extensibility of cell walls in alfalfa (Medicago sativa)

Pectins are abundant in the primary cell walls and involve in regulating intercellular adhesion. MsPG1gene was cloned from alfalfa and its function of Al tolerance was tested in vivo and vitro. Results showed that the expression of MsPG1 was increased by Al, and the MsPG1 was localized in the plasma membrane of epidermal cells in root apexes. Overexpression of MsPG1 in hairy roots of alfalfa increased root growth in the present of Al, which was attributed to the decreased Al accumulations in pectins, hemicellulose 1 (HC1), cell walls and root apexes conferred by the decreased content of HC1 and ratios of negatively charged functional groups (−OH and −COOH) in cell wall. The decreased Al accumulations in pectins and cell walls were also observed in incubation MsPG1 with pectins and cell walls in vitro. TEM and FESEM assays showed that overexpression of MsPG1 protected the cell microstructure of root apexes, and increased the porosity and pore size of porous lattice-like structure of cell wall in the present of Al. These results indicated that MsPG1 could increase Al tolerance of alfalfa via decreasing Al accumulation in cell wall and increasing the porosity and extensibility of cell wall under excess Al stress.

A wheat NBS-LRR gene TaRGA19 participates in Lr19-mediated resistance to Puccinia triticina

Wheat leaf rust, caused by Puccinia triticina (Pt), is one of the most severe fungal diseases on wheat globally. Rational utilization of wheat leaf rust resistance (Lr) genes is still the best choice for control this disease. Wheat seedlings carrying Lr19 showed a high resistance phenotype to all Pt races in China. So far, all the cloned seedling Lr genes including Lr1, Lr10 and Lr21, encode protein with NBS-LRR domain. In this study, a wheat gene with NBS-LRR domain from previously established Lr19-resistance-related cDNA library was cloned and designated as TaRGA19. Full length of this gene was amplified by rapid amplification of cDNA ends (RACE). By blast against IWGSC wheat genome database, we have noticed that TaRGA19 was located on chromosome 2DS, which was different from Lr19 located on chromosome 7DL. Compared with susceptible Thatcher line, expression level of TaRGA19 was upregulated in wheat isogenic lines carrying Lr19 (TcLr19) after inoculation of Pt race THTS. By particle bombardment, TaRGA19-GFP fused protein was localized on plasma membrane of epidermal cells. Using virus-induced gene silencing (VIGS), TaRGA19-knockdown plants of TcLr19 showed reduced resistance and few sporulation phenotype upon Pt challenge. Further histological observation indicated that Pt hyphal growth at the infection sites was less suppressed in the TaRGA19-knockdown plants. In conclusion, we speculate this TaRGA19 gene was involved in the Lr19-mediated resistance to wheat leaf rust along with other components.

Arabidopsis ABCG34 contributes to defense against necrotrophic pathogens by mediating the secretion of camalexin

Plant pathogens cause huge yield losses. Plant defense often depends on toxic secondary metabolites that inhibit pathogen growth. Because most secondary metabolites are also toxic to the plant, specific transporters are needed to deliver them to the pathogens. To identify the transporters that function in plant defense, we screened Arabidopsis thaliana mutants of full-size ABCG transporters for hypersensitivity to sclareol, an antifungal compound. We found that atabcg34 mutants were hypersensitive to sclareol and to the necrotrophic fungi Alternaria brassicicola and Botrytis cinereaAtABCG34 expression was induced by Abrassicicola inoculation as well as by methyl-jasmonate, a defense-related phytohormone, and AtABCG34 was polarly localized at the external face of the plasma membrane of epidermal cells of leaves and roots. atabcg34 mutants secreted less camalexin, a major phytoalexin in Athaliana, whereas plants overexpressing AtABCG34 secreted more camalexin to the leaf surface and were more resistant to the pathogen. When treated with exogenous camalexin, atabcg34 mutants exhibited hypersensitivity, whereas BY2 cells expressing AtABCG34 exhibited improved resistance. Analyses of natural Arabidopsis accessions revealed that AtABCG34 contributes to the disease resistance in naturally occurring genetic variants, albeit to a small extent. Together, our data suggest that AtABCG34 mediates camalexin secretion to the leaf surface and thereby prevents Abrassicicola infection.

RAB-5- and DYNAMIN-1-Mediated Endocytosis of EFF-1 Fusogen Controls Cell-Cell Fusion.

SummaryCell-cell fusion plays essential roles during fertilization and organogenesis. Previous studies in C. elegans led to the identification of the eukaryotic fusion protein (EFF-1 fusogen), which has structural homology to class II viral fusogens. Transcriptional repression of EFF-1 ensures correct fusion fates, and overexpression of EFF-1 results in embryonic lethality. EFF-1 must be expressed on the surface of both fusing cells; however, little is known regarding how cells regulate EFF-1 surface exposure. Here, we report that EFF-1 is actively removed from the plasma membrane of epidermal cells by dynamin- and RAB-5-dependent endocytosis and accumulates in early endosomes. EFF-1 was transiently localized to apical domains of fusion-competent cells. Effective cell-cell fusion occurred only between pairs of cell membranes in which EFF-1 localized. Downregulation of dynamin or RAB-5 caused EFF-1 mislocalization to all apical membrane domains and excessive fusion. Thus, internalization of EFF-1 is a safety mechanism preventing excessive cell fusion.

Simple preparation of plant epidermal tissue for laser microdissection and downstream quantitative proteome and carbohydrate analysis

The outwardly directed cell wall and associated plasma membrane of epidermal cells represent the first layers of plant defense against intruding pathogens. Cell wall modifications and the formation of defense structures at sites of attempted pathogen penetration are decisive for plant defense. A precise isolation of these stress-induced structures would allow a specific analysis of regulatory mechanism and cell wall adaption. However, methods for large-scale epidermal tissue preparation from the model plant Arabidopsis thaliana, which would allow proteome and cell wall analysis of complete, laser-microdissected epidermal defense structures, have not been provided. We developed the adhesive tape – liquid cover glass technique (ACT) for simple leaf epidermis preparation from A. thaliana, which is also applicable on grass leaves. This method is compatible with subsequent staining techniques to visualize stress-related cell wall structures, which were precisely isolated from the epidermal tissue layer by laser microdissection (LM) coupled to laser pressure catapulting. We successfully demonstrated that these specific epidermal tissue samples could be used for quantitative downstream proteome and cell wall analysis. The development of the ACT for simple leaf epidermis preparation and the compatibility to LM and downstream quantitative analysis opens new possibilities in the precise examination of stress- and pathogen-related cell wall structures in epidermal cells. Because the developed tissue processing is also applicable on A. thaliana, well-established, model pathosystems that include the interaction with powdery mildews can be studied to determine principal regulatory mechanisms in plant–microbe interaction with their potential outreach into crop breeding.

Subcellular Redistribution of Root Aquaporins Induced by Hydrogen Peroxide

Aquaporins are water channel proteins that mediate the fine-tuning of cell membrane water permeability during development or in response to environmental stresses. The present work focuses on the oxidative stress-induced redistribution of plasma membrane intrinsic protein (PIP) aquaporins from the plasma membrane (PM) to intracellular membranes. This process was investigated in the Arabidopsis root. Sucrose density gradient centrifugation showed that exposure of roots to 0.5 mM H2O2 induces significant depletion in PM fractions of several abundant PIP homologs after 15 min. Analyses by single-particle tracking and fluorescence correlative spectroscopy showed that, in the PM of epidermal cells, H2O2 treatment induces an increase in lateral motion and a reduction in the density of a fluorescently tagged form of the prototypal AtPIP2;1 isoform, respectively. Co-expression analyses of AtPIP2;1 with endomembrane markers revealed that H2O2 triggers AtPIP2;1 accumulation in the late endosomal compartments. Life-time analyses established that the high stability of PIPs was maintained under oxidative stress conditions, suggesting that H2O2 triggers a mechanism for intracellular sequestration of PM aquaporins without further degradation. In addition to information on cellular regulation of aquaporins, this study provides novel and complementary insights into the dynamic remodeling of plant internal membranes during oxidative stress responses.

Identification of wheat non-specific lipid transfer proteins involved in chilling tolerance.

Three TaLTPs were found to enhance chilling tolerance of transgenic Arabidopsis, which were characterized by analyzes of promoter-GUS activity, subcellular localization, chromosomal location and transcriptional profile. Non-specific lipid transfer proteins (nsLTP) are abundantly expressed in plants, however, their functions are still unclear. In this study, we primarily characterized the functions of 3 type I TaLTP genes that were localized on chromosomes 3A, 3B, and 5D, respectively. The transcripts of TaLTPIb.1 and TaLTPIb.5 were induced under chilling, wound, and drought conditions, while TaLTPId.1 was only up-regulated by dark treatment. All the 3 TaLTP genes could be stimulated by the in vitro treatment of salicylic acid, while TaLTPId.1 was also positively regulated by methyljasmonic acid. Furthermore, the promoter-reporter assay of TaLTPIb.1 in the transgenic brachypodium showed a typical epidermis-specific expression pattern of this gene cluster. When fused with EGFP, all the 3 proteins were shown to localize on the plasma membrane in transgenic tobacco, although a signal in chloroplasts was also observed for TaLTPId.1. Heterogeneous overexpression of each of the TaLTP genes in Arabidopsis resulted in longer root length compared with wild type plants under chilling condition. These results suggest that type I TaLTPs may have a conserved functionality in chilling tolerance by lipid permeation in the plasma membrane of epidermal cells. On the other hand, the type I TaLTPs may exert functional divergence mainly through regulatory subfunctionalization.

Strigolactone analog GR24 triggers changes in PIN2 polarity, vesicle trafficking and actin filament architecture

Strigolactones (SLs) are plant hormones that regulate shoot and root development in a MAX2-dependent manner. The mechanism underlying SLs' effects on roots is unclear. We used root hair elongation to measure root response to SLs. We examined the effects of GR24 (a synthetic, biologically active SL analog) on localization of the auxin efflux transporter PIN2, endosomal trafficking, and F-actin architecture and dynamics in the plasma membrane (PM) of epidermal cells of the primary root elongation zone in wildtype (WT) Arabidopsis and the SL-insensitive mutant max2. We also recorded the response to GR24 of trafficking (tir3), actin (der1) and PIN2 (eir1) mutants. GR24 increased polar localization of PIN2 in the PM of epidermal cells and accumulation of PIN2-containing brefeldin A (BFA) bodies, increased ARA7-labeled endosomal trafficking, reduced F-actin bundling and enhanced actin dynamics, all in a MAX2-dependent manner. Most of the der1 and tir3 mutant lines also displayed reduced sensitivity to GR24 with respect to root hair elongation. We suggest that SLs increase PIN2 polar localization, PIN2 endocytosis, endosomal trafficking, actin debundling and actin dynamics in a MAX2-dependent fashion. This enhancement might underlie the WT root's response to SLs, and suggests noncell autonomous activity of SLs in roots.

Molting and cuticle deposition in the subterranean trichoniscid Titanethes albus (Crustacea, Isopoda)

Terrestrial isopods are a suitable group for the study of cuticle synthesis and calcium dynamics because they molt frequently and have evolved means to store calcium during molt. Little data is currently available on molting in Synocheta and subterranean isopods. We studied the molting dynamics in the subterranean trichoniscid Titanethes albus under laboratory conditions and performed a microscopic investigation of sternal CaCO3 deposits and the tergal epithelium during molt in this species. In accordance with its lower metabolic rate, molting in the laboratory is roughly 2–3 times less frequent in Titanethes albus than would be expected for an epigean isopod under similar conditions. Animals assumed characteristic postures following the molt of each body half and did not consume the posterior exuviae after posterior molt. The structure of sternal calcium deposits and the ultrastructural characteristics of the epidermis during cuticle formation in Titanethes albus are similar to those described in representatives of Ligiidae. During the deposition of the exocuticle, the apical plasma membrane of epidermal cells forms finger-like extensions and numerous invaginations. In the ecdysial space of individuals in late premolt we observed cellular extensions surrounded by bundles of tubules.

A member of the PLEIOTROPIC DRUG RESISTANCE family of ATP binding cassette transporters is required for the formation of a functional cuticle in Arabidopsis.

Although the multilayered structure of the plant cuticle was discovered many years ago, the molecular basis of its formation and the functional relevance of the layers are not understood. Here, we present the permeable cuticle1 (pec1) mutant of Arabidopsis thaliana, which displays features associated with a highly permeable cuticle in several organs. In pec1 flowers, typical cutin monomers, such as ω-hydroxylated fatty acids and 10,16-dihydroxypalmitate, are reduced to 40% of wild-type levels and are accompanied by the appearance of lipidic inclusions within the epidermal cell. The cuticular layer of the cell wall, rather than the cuticle proper, is structurally altered in pec1 petals. Therefore, a significant role for the formation of the diffusion barrier in petals can be attributed to this layer. Thus, pec1 defines a new class of mutants. The phenotypes of the pec1 mutant are caused by the knockout of ATP BINDING CASSETTEG32 (ABCG32), an ABC transporter from the PLEIOTROPIC DRUG RESISTANCE family that is localized at the plasma membrane of epidermal cells in a polar manner toward the surface of the organs. Our results suggest that ABCG32 is involved in the formation of the cuticular layer of the cell wall, most likely by exporting particular cutin precursors from the epidermal cell.

Mapping and functional analysis of four apple receptor-like protein kinases related to LRPKm1 in HcrVf2-transgenic and wild-type apple plants

The Malus–Venturia inaequalis interaction is the most studied plant–pathogen interaction involving a woody species. Besides the cloning of an apple scab resistance gene HcrVf2, several sequences have been recently identified that are modulated after pathogen recognition in Vf-resistant genotypes. Among these, there is a putative leucine-rich repeat receptor-like protein kinase from the apple scab-resistant cv. Florina, named LRPKm1 that is induced after V. inaequalis inoculation and salicylic acid treatment. In this work, the isolation, characterization, and mapping of four new genes belonging to the LRPKm multigene family are reported. According to their cumulative expression profiles in HcrVf2-transgenic and wild-type apple plants treated with V. inaequalis, LRPKm genes have been divided in two groups. LRPKm1 and LRPKm3, giving a response related to the presence of HcrVf2, are probably involved in the recognition of pathogen-derived signals. LRPKm2 and LRPKm4, with an expression profile unrelated to the HcrVf2 gene, are putatively involved in the plant basal defense. Furthermore, we have localized LRPKm proteins at the cytological level in the plasma membrane of epidermal cells in resistant genotypes following pathogen challenge, thus confirming software predictions and molecular results. The possible involvement of LRPKm proteins in apple scab resistance and in the plant basal defense makes them attractive for a better comprehension of the molecular mechanisms of the signal transduction pathways after pathogen recognition.

ABC-type transporters and cuticle assembly

The aerial organs of plants are covered with a cuticle, a continuous layer overlaying the outermost cell walls of the epidermis. The cuticle is composed of two major classes of the lipid biopolymers: cutin and waxes, collectively termed cuticular lipids. Biosynthesis and transport of cuticular lipids occur predominantly in the epidermis cells. In the transport pathway, cuticular lipids are exported from their site of biosynthesis in the ER/plastid to the extracellular space through the plasma membrane and cell wall. Growing evidence suggests that ATP-binding cassette (ABC) transporters are implicated in transport of cuticular lipids across the plasma membrane of epidermal cells. The Arabidopsis ABC-type transporter protein CER5 (WBC12) was reported to act as a wax monomer transporter. In recent works, our group and others showed that a CER5-related protein, DESPERADO (DSO/WBC11), is required for cutin and wax monomers transport through the plasma membrane of Arabidopsis epidermis cells. Unlike the cer5 mutant, DSO loss-of-function had a profound effect on plant growth and development, particularly dwarfism, postgenital organ fusions, and altered epidermal cell differentiation. The partially overlapping function of CER5 and DSO and the fact that these proteins are half-size ABC transporters suggest that they might form a hetero-dimeric complex while transporting wax components. An intriguing observation was the polar localization of DSO in the distal part of epidermis cells. This polar expression might be explained by DSO localization within lipid rafts, specific plasma membrane microdomains which are associated with polar protein expression. In this review we suggest possible mechanisms for cuticular lipids transport and a link between DSO function and polar expression. Furthermore, we also discuss the subsequent transport of cuticular constituents through the hydrophobic cell wall and the possible involvement of lipid transfer proteins in this process.Addendum to: Panikashvili D, Savaldi-Goldstein S, Mandel T, Yifhar T, Franke RB, Höfer R, Schreiber L, Chory J, Aharoni A. The Arabidopsis DESPERADO/AtWBC11 transporter is required for cutin and wax secretion. Plant Physiol 2007; 145:1345-60.

Assembly of the Drosophila larval exoskeleton requires controlled secretion and shaping of the apical plasma membrane

The apical plasma membrane of epithelial cells plays a central role in producing and shaping the apical extracellular matrix (aECM) that eventually adopts a stereotypic architecture required for the physical and physiological needs of the epithelium. To assess the implication of the apical plasma membrane on aECM differentiation, we have studied the function of the apical plasma membrane t-SNARE Syntaxin 1A in the embryo of the fruit fly Drosophila melanogaster during differentiation of the stratified exoskeleton, the cuticle, which is composed of proteins and the polysaccharide chitin. The cuticle layers of syntaxin1A deficient larvae are rudimentary. Consistently, Syntaxin 1A is required for the secretion of O-glycosylated proteins and components involved in pigmentation and protein cross-linking. By contrast, localization of chitin synthesis and organising proteins to the apical plasma membrane or to the extracellular space does not depend on Syntaxin 1A activity. However, chitin microfibrils have a random orientation instead of being arranged in parallel. This correlates with the lack of corrugations at the apical plasma membrane of epidermal cells, the apical undulae that have been proposed to be crucial for chitin microfibril orientation. Hence, Syntaxin 1A contributes to cuticle differentiation by controlling correct apical plasma membrane topology as well as mediating secretion of a subset of extracellular proteins required for layer organisation. Our data also indicate that yet another unidentified t-SNARE is needed in parallel to Syntaxin 1A to deliver extracellular material for complete cuticle assembly. Evidently, coordination of apical membrane modelling and two secretion routes are essential for stereotypic aECM organisation.

Repression of Cell-Cell Fusion by Components of the C. elegans Vacuolar ATPase Complex

Cell-cell fusion initiates fertilization, sculpts tissues during animal development, reprograms stem cells to new differentiated states, and may be a key step in cancer progression. While cell fusion is tightly regulated, the mechanisms that limit fusion to appropriate partners are unknown. Here, we report that the fus-1 gene is essential to repress fusion of epidermal cells in C. elegans: in severe fus-1 mutants, all epidermal cells, except the lateral seam cells, inappropriately fuse into a single large syncytium. This hyperfusion requires EFF-1, an integral membrane protein essential for fusion of epidermal cells into discrete syncytia. FUS-1 is localized to the apical plasma membrane in all epidermal cells potentiated to undergo fusion, whereas it is virtually undetectable in nonfusing seam cells. fus-1 encodes the e subunit of the vacuolar H(+)-ATPase (V-ATPase), and loss of other V-ATPase subunits also causes widespread hyperfusion. These findings raise the possibility of manipulating cell fusion by altering V-ATPase activity.

Plant Cuticular Lipid Export Requires an ABC Transporter

A waxy protective cuticle coats all primary aerial plant tissues. Its synthesis requires extensive export of lipids from epidermal cells to the plant surface. Arabidopsis cer5 mutants had reduced stem cuticular wax loads and accumulated sheetlike inclusions in the cytoplasm of wax-secreting cells. These inclusions represented abnormal deposits of cuticular wax and resembled inclusions found in a human disorder caused by a defective peroxisomal adenosine triphosphate binding cassette (ABC) transporter. We found that the CER5 gene encodes an ABC transporter localized in the plasma membrane of epidermal cells and conclude that it is required for wax export to the cuticle.

Integrin-like protein at the invaginated plasma membrane of epidermal cells in mature leaves of the marine angiosperm Zostera marina L.

By immunoblotting with anti-human integrin beta polyclonal antibodies (beta1, beta3 or beta5), a single distinct band of about 60 kDa was detected in total protein extracts from mature leaves of the seagrass Zostera marina L., but no band was detected in total protein extracts from immature seagrass leaves, freshwater grass leaves or Arabidopsis thaliana (L.) Heynh. leaves. This integrin-like protein was detected by indirect immunofluorescence microscopy on the surface of non-spherical protoplasts of epidermal cells isolated from mature seagrass leaves using an anti-integrin beta3 polyclonal antibody. Electron-microscopic analyses with the same antibody indicated that this integrin-like protein was localized specifically in the invaginated plasma membrane of epidermal cells in mature seagrass leaves. Therefore, this integrin-like protein of about 60 kDa may be involved in the developmentally regulated invagination of the plasma membrane in epidermal cells of the seagrass Z. marina.

New cellulose synthesizing complexes (terminal complexes) involved in animal cellulose biosynthesis in the tunicateMetandrocarpa uedai

Cellulose synthesizing enzyme complexes (terminal complexes, TCs) have been found in the plasma membrane of epidermal cells in the tunicateMetandrocarpa uedai by using freeze-fracture replication techniques for electron microscopy. Assembly of cellulose microfibrils by TCs is a universal phenomenon in the biological kingdoms. The TCs are locally distributed in the plasma membrane of the epidermal cells facing the tunic, and no TCs are observed on the lateral membranes bordered by tight junctions. The TCs consist of two types of membrane subunits: large particles (14.5 nm in diameter) on the periphery and small subunit particles (7.2 nm) filling the center; the latter are hypothesized to be involved in cellulose synthesis. The TCs are the linear type (ca. 195 nm in length and 78 nm in width). Direct connections of TCs with the termini of microfibrils were observed. Amorphous regions, which were hypothesized the nascent microfibrils, were associated with the depressions of the TCs. The distortion of microfibrils on their terminus indicates that the crystallization may occur at the margin of TCs from which the microfibrils are discharged. This report provides evidence that: (1) The outer cell membrane of epidermis is the site for the assembly of cellulose microfibrils in the tunic; (2) a new type of TC is involved in the biosynthesis of cellulose microfibrils in the tunicates; (3) disorganized glucan chains may be synthesized in the depression of TCs and crystallized outside the E-surface of the epidermal cell membrane.

The effect of brefeldin A on the redistribution of osmiophilic particles and the gravitropic response of rye coleoptiles

The occurrence of IAA-inducible osmiophilic particles (OPs) in the periplasmic space of epidermal cells in the upper and lower flank (UF, LF) of gravistimulated rye coleoptile segments was analyzed employing brefeldin A (BFA) as an inhibitor of secretion at the plasma membrane. A 2 h horizontal gravistimulation of untreated samples caused a duplication of OPs in the periplasmic space of epidermal cells at the growth-inhibited UF as compared to the LF of upward bending coleoptile segments. In contrast to this, the number of OPs within the cytoplasm close to the plasma membrane of epidermal cells was similar at both flanks. BFA caused an inhibition of graviresponsive growth and prevented the occurrence of OPs in the periplasmic space of the epidermal cells of the UF and the LF. Likewise, growth of vertically oriented coleoptile segments was inhibited by BFA. Growth inhibition of both gravistimulated and control segments was accompanied by a twofold increase of the occurrence of cytoplasmic OPs. The results illustrate that the occurrence of OPs within the periplasmic space of the epidermal cells depends on secretion processes. Furthermore they provide evidence that their increased occurrence in the growth-inhibited UF during gravistimulation is due to their inhibited infiltration into the cell walls. We suggest that thereby wall loosening is temporarily prevented.