Changes in Cell Wall Properties Coincide with Overexpression of Extensin Fusion Proteins in Suspension Cultured Tobacco Cells

DOE PAGES

Tan, Li; Pu, Yunqiao; Pattathil, Sivakumar; ...

2014-12-23

Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp4 glycosylation motifs and intermolecular cross-linking motifs such as the TyrXaaTyr sequence. Extensins are believed to form a cross-linked network in the plant cell wall through the tyrosine-derivatives isodityrosine, pulcherosine, and di-isodityrosine. Overexpression of three synthetic genes encoding different elastin-arabinogalactan protein-extensin hybrids in tobacco suspension cultured cells yielded novel cross-linking glycoproteins that shared features of the extensins, arabinogalactan proteins and elastin. The cell wall properties of the three transgenic cell lines were all changed, but in different ways. One transgenic cell line showed decreased cellulose crystallinity and increasedmore » wall xyloglucan content; the second transgenic cell line contained dramatically increased hydration capacity and notably increased cell wall biomass, increased di-isodityrosine, and increased protein content; the third transgenic cell line displayed wall phenotypes similar to wild type cells, except changed xyloglucan epitope extractability. In conclusion, these data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.« less

Network reconstruction and systems analysis of plant cell wall deconstruction by Neurospora crassa.

PubMed

Samal, Areejit; Craig, James P; Coradetti, Samuel T; Benz, J Philipp; Eddy, James A; Price, Nathan D; Glass, N Louise

2017-01-01

Plant biomass degradation by fungal-derived enzymes is rapidly expanding in economic importance as a clean and efficient source for biofuels. The ability to rationally engineer filamentous fungi would facilitate biotechnological applications for degradation of plant cell wall polysaccharides. However, incomplete knowledge of biomolecular networks responsible for plant cell wall deconstruction impedes experimental efforts in this direction. To expand this knowledge base, a detailed network of reactions important for deconstruction of plant cell wall polysaccharides into simple sugars was constructed for the filamentous fungus Neurospora crassa . To reconstruct this network, information was integrated from five heterogeneous data types: functional genomics, transcriptomics, proteomics, genetics, and biochemical characterizations. The combined information was encapsulated into a feature matrix and the evidence weighted to assign annotation confidence scores for each gene within the network. Comparative analyses of RNA-seq and ChIP-seq data shed light on the regulation of the plant cell wall degradation network, leading to a novel hypothesis for degradation of the hemicellulose mannan. The transcription factor CLR-2 was subsequently experimentally shown to play a key role in the mannan degradation pathway of N. crassa . Here we built a network that serves as a scaffold for integration of diverse experimental datasets. This approach led to the elucidation of regulatory design principles for plant cell wall deconstruction by filamentous fungi and a novel function for the transcription factor CLR-2. This expanding network will aid in efforts to rationally engineer industrially relevant hyper-production strains.

Identifying new lignin bioengineering targets: 1. Monolignol-substitute impacts on lignin formation and cell wall fermentability

PubMed Central

2010-01-01

Background Recent discoveries highlighting the metabolic malleability of plant lignification indicate that lignin can be engineered to dramatically alter its composition and properties. Current plant biotechnology efforts are primarily aimed at manipulating the biosynthesis of normal monolignols, but in the future apoplastic targeting of phenolics from other metabolic pathways may provide new approaches for designing lignins that are less inhibitory toward the enzymatic hydrolysis of structural polysaccharides, both with and without biomass pretreatment. To identify promising new avenues for lignin bioengineering, we artificially lignified cell walls from maize cell suspensions with various combinations of normal monolignols (coniferyl and sinapyl alcohols) plus a variety of phenolic monolignol substitutes. Cell walls were then incubated in vitro with anaerobic rumen microflora to assess the potential impact of lignin modifications on the enzymatic degradability of fibrous crops used for ruminant livestock or biofuel production. Results In the absence of anatomical constraints to digestion, lignification with normal monolignols hindered both the rate and extent of cell wall hydrolysis by rumen microflora. Inclusion of methyl caffeate, caffeoylquinic acid, or feruloylquinic acid with monolignols considerably depressed lignin formation and strikingly improved the degradability of cell walls. In contrast, dihydroconiferyl alcohol, guaiacyl glycerol, epicatechin, epigallocatechin, and epigallocatechin gallate readily formed copolymer-lignins with normal monolignols; cell wall degradability was moderately enhanced by greater hydroxylation or 1,2,3-triol functionality. Mono- or diferuloyl esters with various aliphatic or polyol groups readily copolymerized with monolignols, but in some cases they accelerated inactivation of wall-bound peroxidase and reduced lignification; cell wall degradability was influenced by lignin content and the degree of ester group hydroxylation. Conclusion Overall, monolignol substitutes improved the inherent degradability of non-pretreated cell walls by restricting lignification or possibly by reducing lignin hydrophobicity or cross-linking to structural polysaccharides. Furthermore some monolignol substitutes, chiefly readily cleaved bi-phenolic conjugates like epigallocatechin gallate or diferuloyl polyol esters, are expected to greatly boost the enzymatic degradability of cell walls following chemical pretreatment. In ongoing work, we are characterizing the enzymatic saccharification of intact and chemically pretreated cell walls lignified by these and other monolignol substitutes to identify promising genetic engineering targets for improving plant fiber utilization. PMID:20565789

Expression of S-adenosylmethionine Hydrolase in Tissues Synthesizing Secondary Cell Walls Alters Specific Methylated Cell Wall Fractions and Improves Biomass Digestibility

DOE PAGES

Eudes, Aymerick; Zhao, Nanxia; Sathitsuksanoh, Noppadon; ...

2016-07-19

Plant biomass is a large source of fermentable sugars for the synthesis of bioproducts using engineered microbes. These sugars are stored as cell wall polymers, mainly cellulose and hemicellulose, and are embedded with lignin, which makes their enzymatic hydrolysis challenging. One of the strategies to reduce cell wall recalcitrance is the modification of lignin content and composition. Lignin is a phenolic polymer of methylated aromatic alcohols and its synthesis in tissues developing secondary cell walls is a significant sink for the consumption of the methyl donor S-adenosylmethionine (AdoMet). In this study, we demonstrate in Arabidopsis stems that targeted expression ofmore » AdoMet hydrolase (AdoMetase, E.C. 3.3.1.2) in secondary cell wall synthesizing tissues reduces the AdoMet pool and impacts lignin content and composition. In particular, both NMR analysis and pyrolysis gas chromatography mass spectrometry of lignin in engineered biomass showed relative enrichment of non-methylated p-hydroxycinnamyl (H) units and a reduction of dimethylated syringyl (S) units. This indicates a lower degree of methylation compared to that in wild-type lignin. Quantification of cell wall-bound hydroxycinnamates revealed a reduction of ferulate in AdoMetase transgenic lines. Biomass from transgenic lines, in contrast to that in control plants, exhibits an enrichment of glucose content and a reduction in the degree of hemicellulose glucuronoxylan methylation. We also show that these modifications resulted in a reduction of cell wall recalcitrance, because sugar yield generated by enzymatic biomass saccharification was greater than that of wild-type plants. Considering that transgenic plants show no important diminution of biomass yields, and that heterologous expression of AdoMetase protein can be spatiotemporally optimized, this novel approach provides a valuable option for the improvement of lignocellulosic biomass feedstock.« less

Expression of S-adenosylmethionine Hydrolase in Tissues Synthesizing Secondary Cell Walls Alters Specific Methylated Cell Wall Fractions and Improves Biomass Digestibility

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eudes, Aymerick; Zhao, Nanxia; Sathitsuksanoh, Noppadon

Plant biomass is a large source of fermentable sugars for the synthesis of bioproducts using engineered microbes. These sugars are stored as cell wall polymers, mainly cellulose and hemicellulose, and are embedded with lignin, which makes their enzymatic hydrolysis challenging. One of the strategies to reduce cell wall recalcitrance is the modification of lignin content and composition. Lignin is a phenolic polymer of methylated aromatic alcohols and its synthesis in tissues developing secondary cell walls is a significant sink for the consumption of the methyl donor S-adenosylmethionine (AdoMet). In this study, we demonstrate in Arabidopsis stems that targeted expression ofmore » AdoMet hydrolase (AdoMetase, E.C. 3.3.1.2) in secondary cell wall synthesizing tissues reduces the AdoMet pool and impacts lignin content and composition. In particular, both NMR analysis and pyrolysis gas chromatography mass spectrometry of lignin in engineered biomass showed relative enrichment of non-methylated p-hydroxycinnamyl (H) units and a reduction of dimethylated syringyl (S) units. This indicates a lower degree of methylation compared to that in wild-type lignin. Quantification of cell wall-bound hydroxycinnamates revealed a reduction of ferulate in AdoMetase transgenic lines. Biomass from transgenic lines, in contrast to that in control plants, exhibits an enrichment of glucose content and a reduction in the degree of hemicellulose glucuronoxylan methylation. We also show that these modifications resulted in a reduction of cell wall recalcitrance, because sugar yield generated by enzymatic biomass saccharification was greater than that of wild-type plants. Considering that transgenic plants show no important diminution of biomass yields, and that heterologous expression of AdoMetase protein can be spatiotemporally optimized, this novel approach provides a valuable option for the improvement of lignocellulosic biomass feedstock.« less

Enhancing cellulose utilization for fuels and chemicals by genetic modification of plant cell wall architecture.

PubMed

Vermerris, Wilfred; Abril, Alejandra

2015-04-01

Cellulose from plant biomass can serve as a sustainable feedstock for fuels, chemicals and polymers that are currently produced from petroleum. In order to enhance economic feasibility, the efficiency of cell wall deconstruction needs to be enhanced. With the use of genetic and biotechnological approaches cell wall composition can be modified in such a way that interactions between the major cell wall polymers—cellulose, hemicellulosic polysaccharides and lignin—are altered. Some of the resulting plants are compromised in their growth and development, but this may be caused in part by the plant's overcompensation for metabolic perturbances. In other cases novel structures have been introduced in the cell wall without negative effects. The first field studies with engineered bioenergy crops look promising, while detailed structural analyses of cellulose synthase offer new opportunities to modify cellulose itself. Copyright © 2014 Elsevier Ltd. All rights reserved.

Arterial grafts exhibiting unprecedented cellular infiltration and remodeling in vivo: the role of cells in the vascular wall.

PubMed

Row, Sindhu; Peng, Haofan; Schlaich, Evan M; Koenigsknecht, Carmon; Andreadis, Stelios T; Swartz, Daniel D

2015-05-01

To engineer and implant vascular grafts in the arterial circulation of a pre-clinical animal model and assess the role of donor medial cells in graft remodeling and function. Vascular grafts were engineered using Small Intestinal Submucosa (SIS)-fibrin hybrid scaffold and implanted interpositionally into the arterial circulation of an ovine model. We sought to demonstrate implantability of SIS-Fibrin based grafts; examine the remodeling; and determine whether the presence of vascular cells in the medial wall was necessary for cellular infiltration from the host and successful remodeling of the implants. We observed no occlusions or anastomotic complications in 18 animals that received these grafts. Notably, the grafts exhibited unprecedented levels of host cell infiltration that was not limited to the anastomotic sites but occurred through the lumen as well as the extramural side, leading to uniform cell distribution. Incoming cells remodeled the extracellular matrix and matured into functional smooth muscle cells as evidenced by expression of myogenic markers and development of vascular reactivity. Interestingly, tracking the donor cells revealed that their presence was beneficial but not necessary for successful grafting. Indeed, the proliferation rate and number of donor cells decreased over time as the vascular wall was dominated by host cells leading to significant remodeling and development of contractile function. These results demonstrate that SIS-Fibrin grafts can be successfully implanted into the arterial circulation of a clinically relevant animal model, improve our understanding of vascular graft remodeling and raise the possibility of engineering mural cell-free arterial grafts. Copyright © 2015 Elsevier Ltd. All rights reserved.

Multiple-Step Injection Molding for Fibrin-Based Tissue-Engineered Heart Valves

PubMed Central

Weber, Miriam; Gonzalez de Torre, Israel; Moreira, Ricardo; Frese, Julia; Oedekoven, Caroline; Alonso, Matilde; Rodriguez Cabello, Carlos J.

2015-01-01

Heart valves are elaborate and highly heterogeneous structures of the circulatory system. Despite the well accepted relationship between the structural and mechanical anisotropy and the optimal function of the valves, most approaches to create tissue-engineered heart valves (TEHVs) do not try to mimic this complexity and rely on one homogenous combination of cells and materials for the whole construct. The aim of this study was to establish an easy and versatile method to introduce spatial diversity into a heart valve fibrin scaffold. We developed a multiple-step injection molding process that enables the fabrication of TEHVs with heterogeneous composition (cell/scaffold material) of wall and leaflets without the need of gluing or suturing components together, with the leaflets firmly connected to the wall. The integrity of the valves and their functionality was proved by either opening/closing cycles in a bioreactor (proof of principle without cells) or with continuous stimulation over 2 weeks. We demonstrated the potential of the method by the two-step molding of the wall and the leaflets containing different cell lines. Immunohistology after stimulation confirmed tissue formation and demonstrated the localization of the different cell types. Furthermore, we showed the proof of principle fabrication of valves using different materials for wall (fibrin) and leaflets (hybrid gel of fibrin/elastin-like recombinamer) and with layered leaflets. The method is easy to implement, does not require special facilities, and can be reproduced in any tissue-engineering lab. While it has been demonstrated here with fibrin, it can easily be extended to other hydrogels. PMID:25654448

Multiple-Step Injection Molding for Fibrin-Based Tissue-Engineered Heart Valves.

PubMed

Weber, Miriam; Gonzalez de Torre, Israel; Moreira, Ricardo; Frese, Julia; Oedekoven, Caroline; Alonso, Matilde; Rodriguez Cabello, Carlos J; Jockenhoevel, Stefan; Mela, Petra

2015-08-01

Heart valves are elaborate and highly heterogeneous structures of the circulatory system. Despite the well accepted relationship between the structural and mechanical anisotropy and the optimal function of the valves, most approaches to create tissue-engineered heart valves (TEHVs) do not try to mimic this complexity and rely on one homogenous combination of cells and materials for the whole construct. The aim of this study was to establish an easy and versatile method to introduce spatial diversity into a heart valve fibrin scaffold. We developed a multiple-step injection molding process that enables the fabrication of TEHVs with heterogeneous composition (cell/scaffold material) of wall and leaflets without the need of gluing or suturing components together, with the leaflets firmly connected to the wall. The integrity of the valves and their functionality was proved by either opening/closing cycles in a bioreactor (proof of principle without cells) or with continuous stimulation over 2 weeks. We demonstrated the potential of the method by the two-step molding of the wall and the leaflets containing different cell lines. Immunohistology after stimulation confirmed tissue formation and demonstrated the localization of the different cell types. Furthermore, we showed the proof of principle fabrication of valves using different materials for wall (fibrin) and leaflets (hybrid gel of fibrin/elastin-like recombinamer) and with layered leaflets. The method is easy to implement, does not require special facilities, and can be reproduced in any tissue-engineering lab. While it has been demonstrated here with fibrin, it can easily be extended to other hydrogels.

Identifying the ionically bound cell wall and intracellular glycoside hydrolases in late growth stage Arabidopsis stems: implications for the genetic engineering of bioenergy crops.

PubMed

Wei, Hui; Brunecky, Roman; Donohoe, Bryon S; Ding, Shi-You; Ciesielski, Peter N; Yang, Shihui; Tucker, Melvin P; Himmel, Michael E

2015-01-01

Identifying the cell wall-ionically bound glycoside hydrolases (GHs) in Arabidopsis stems is important for understanding the regulation of cell wall integrity. For cell wall proteomics studies, the preparation of clean cell wall fractions is a challenge since cell walls constitute an open compartment, which is more likely to contain a mixture of intracellular and extracellular proteins due to cell leakage at the late growth stage. Here, we utilize a CaCl2-extraction procedure to isolate non-structural proteins from Arabidopsis whole stems, followed by the in-solution and in-gel digestion methods coupled with Nano-LC-MS/MS, bioinformatics and literature analyses. This has led to the identification of 75 proteins identified using the in-solution method and 236 proteins identified by the in-gel method, among which about 10% of proteins predicted to be secreted. Together, eight cell wall proteins, namely AT1G75040, AT5G26000, AT3G57260, AT4G21650, AT3G52960, AT3G49120, AT5G49360, and AT3G14067, were identified by the in-solution method; among them, three were the GHs (AT5G26000, myrosinase 1, GH1; AT3G57260, β-1,3-glucanase 2, GH17; AT5G49360, bifunctional XYL 1/α-L-arabinofuranosidase, GH3). Moreover, four more GHs: AT4G30270 (xyloglucan endotransferase, GH16), AT1G68560 (bifunctional α-l-arabinofuranosidase/XYL, GH31), AT1G12240 (invertase, GH32) and AT2G28470 (β-galactosidase 8, GH35), were identified by the in-gel solution method only. Notably, more than half of above identified GHs are xylan- or hemicellulose-modifying enzymes, and will likely have an impact on cellulose accessibility, which is a critical factor for downstream enzymatic hydrolysis of plant tissues for biofuels production. The implications of these cell wall proteins identified at the late growth stage for the genetic engineering of bioenergy crops are discussed.

Identifying the ionically bound cell wall and intracellular glycoside hydrolases in late growth stage Arabidopsis stems: Implications for the genetic engineering of bioenergy crops

DOE PAGES

Wei, Hui; Brunecky, Roman; Donohoe, Bryon S.; ...

2015-05-13

Identifying the cell wall-ionically bound glycoside hydrolases (GHs) in Arabidopsis stems is important for understanding the regulation of cell wall integrity. For cell wall proteomics studies, the preparation of clean cell wall fractions is a challenge since cell walls constitute an open compartment, which is more likely to contain a mixture of intracellular and extracellular proteins due to cell leakage at the late growth stage. Here, for this study, we utilize a CaCl 2-extraction procedure to isolate non-structural proteins from Arabidopsis whole stems, followed by the in-solution and in-gel digestion methods coupled with Nano-LC-MS/MS, bioinformatics and literature analyses. This hasmore » led to the identification of 75 proteins identified using the in-solution method and 236 proteins identified by the in-gel method, among which about 10% of proteins predicted to be secreted. Together, eight cell wall proteins, namely AT1G75040, AT5G26000, AT3G57260, AT4G21650, AT3G52960, AT3G49120, AT5G49360, and AT3G14067, were identified by the in-solution method; among them, three were the GHs (AT5G26000, myrosinase 1, GH1; AT3G57260, β-1,3-glucanase 2, GH17; AT5G49360, bifunctional XYL 1/α-L-arabinofuranosidase, GH3). Moreover, four more GHs: AT4G30270 (xyloglucan endotransferase, GH16), AT1G68560 (bifunctional α-l-arabinofuranosidase/XYL, GH31), AT1G12240 (invertase, GH32) and AT2G28470 (β-galactosidase 8, GH35), were identified by the in-gel solution method only. Notably, more than half of above identified GHs are xylan- or hemicellulose-modifying enzymes, and will likely have an impact on cellulose accessibility, which is a critical factor for downstream enzymatic hydrolysis of plant tissues for biofuels production. Finally, the implications of these cell wall proteins identified at the late growth stage for the genetic engineering of bioenergy crops are discussed.« less

Identifying the ionically bound cell wall and intracellular glycoside hydrolases in late growth stage Arabidopsis stems: implications for the genetic engineering of bioenergy crops

PubMed Central

Wei, Hui; Brunecky, Roman; Donohoe, Bryon S.; Ding, Shi-You; Ciesielski, Peter N.; Yang, Shihui; Tucker, Melvin P.; Himmel, Michael E.

2015-01-01

Identifying the cell wall-ionically bound glycoside hydrolases (GHs) in Arabidopsis stems is important for understanding the regulation of cell wall integrity. For cell wall proteomics studies, the preparation of clean cell wall fractions is a challenge since cell walls constitute an open compartment, which is more likely to contain a mixture of intracellular and extracellular proteins due to cell leakage at the late growth stage. Here, we utilize a CaCl2-extraction procedure to isolate non-structural proteins from Arabidopsis whole stems, followed by the in-solution and in-gel digestion methods coupled with Nano-LC-MS/MS, bioinformatics and literature analyses. This has led to the identification of 75 proteins identified using the in-solution method and 236 proteins identified by the in-gel method, among which about 10% of proteins predicted to be secreted. Together, eight cell wall proteins, namely AT1G75040, AT5G26000, AT3G57260, AT4G21650, AT3G52960, AT3G49120, AT5G49360, and AT3G14067, were identified by the in-solution method; among them, three were the GHs (AT5G26000, myrosinase 1, GH1; AT3G57260, β-1,3-glucanase 2, GH17; AT5G49360, bifunctional XYL 1/α-L-arabinofuranosidase, GH3). Moreover, four more GHs: AT4G30270 (xyloglucan endotransferase, GH16), AT1G68560 (bifunctional α-l-arabinofuranosidase/XYL, GH31), AT1G12240 (invertase, GH32) and AT2G28470 (β-galactosidase 8, GH35), were identified by the in-gel solution method only. Notably, more than half of above identified GHs are xylan- or hemicellulose-modifying enzymes, and will likely have an impact on cellulose accessibility, which is a critical factor for downstream enzymatic hydrolysis of plant tissues for biofuels production. The implications of these cell wall proteins identified at the late growth stage for the genetic engineering of bioenergy crops are discussed. PMID:26029221

Identifying the ionically bound cell wall and intracellular glycoside hydrolases in late growth stage Arabidopsis stems: Implications for the genetic engineering of bioenergy crops

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wei, Hui; Brunecky, Roman; Donohoe, Bryon S.

Identifying the cell wall-ionically bound glycoside hydrolases (GHs) in Arabidopsis stems is important for understanding the regulation of cell wall integrity. For cell wall proteomics studies, the preparation of clean cell wall fractions is a challenge since cell walls constitute an open compartment, which is more likely to contain a mixture of intracellular and extracellular proteins due to cell leakage at the late growth stage. Here, for this study, we utilize a CaCl 2-extraction procedure to isolate non-structural proteins from Arabidopsis whole stems, followed by the in-solution and in-gel digestion methods coupled with Nano-LC-MS/MS, bioinformatics and literature analyses. This hasmore » led to the identification of 75 proteins identified using the in-solution method and 236 proteins identified by the in-gel method, among which about 10% of proteins predicted to be secreted. Together, eight cell wall proteins, namely AT1G75040, AT5G26000, AT3G57260, AT4G21650, AT3G52960, AT3G49120, AT5G49360, and AT3G14067, were identified by the in-solution method; among them, three were the GHs (AT5G26000, myrosinase 1, GH1; AT3G57260, β-1,3-glucanase 2, GH17; AT5G49360, bifunctional XYL 1/α-L-arabinofuranosidase, GH3). Moreover, four more GHs: AT4G30270 (xyloglucan endotransferase, GH16), AT1G68560 (bifunctional α-l-arabinofuranosidase/XYL, GH31), AT1G12240 (invertase, GH32) and AT2G28470 (β-galactosidase 8, GH35), were identified by the in-gel solution method only. Notably, more than half of above identified GHs are xylan- or hemicellulose-modifying enzymes, and will likely have an impact on cellulose accessibility, which is a critical factor for downstream enzymatic hydrolysis of plant tissues for biofuels production. Finally, the implications of these cell wall proteins identified at the late growth stage for the genetic engineering of bioenergy crops are discussed.« less

Gene stacking of multiple traits for high yield of fermentable sugars in plant biomass

DOE PAGES

Aznar, Aude; Chalvin, Camille; Shih, Patrick M.; ...

2018-01-09

Second-generation biofuels produced from biomass can help to decrease dependency on fossil fuels, bringing about many economic and environmental benefits. To make biomass more suitable for biorefinery use, we need a better understanding of plant cell wall biosynthesis. Increasing the ratio of C6 to C5 sugars in the cell wall and decreasing the lignin content are two important targets in engineering of plants that are more suitable for downstream processing for second-generation biofuel production. Here, we have studied the basic mechanisms of cell wall biosynthesis and identified genes involved in biosynthesis of pectic galactan, including the GALS1 galactan synthase andmore » the UDP-galactose/UDP-rhamnose transporter URGT1. We have engineered plants with a more suitable biomass composition by applying these findings, in conjunction with synthetic biology and gene stacking tools. Plants were engineered to have up to fourfold more pectic galactan in stems by overexpressing GALS1, URGT1, and UGE2, a UDP-glucose epimerase. Furthermore, the increased galactan trait was engineered into plants that were already engineered to have low xylan content by restricting xylan biosynthesis to vessels where this polysaccharide is essential. Finally, the high galactan and low xylan traits were stacked with the low lignin trait obtained by expressing the QsuB gene encoding dehydroshikimate dehydratase in lignifying cells. In conclusion, the results show that approaches to increasing C6 sugar content, decreasing xylan, and reducing lignin content can be combined in an additive manner. Thus, the engineered lines obtained by this trait-stacking approach have substantially improved properties from the perspective of biofuel production, and they do not show any obvious negative growth effects. The approach used in this study can be readily transferred to bioenergy crop plants.« less

Gene stacking of multiple traits for high yield of fermentable sugars in plant biomass

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aznar, Aude; Chalvin, Camille; Shih, Patrick M.

Second-generation biofuels produced from biomass can help to decrease dependency on fossil fuels, bringing about many economic and environmental benefits. To make biomass more suitable for biorefinery use, we need a better understanding of plant cell wall biosynthesis. Increasing the ratio of C6 to C5 sugars in the cell wall and decreasing the lignin content are two important targets in engineering of plants that are more suitable for downstream processing for second-generation biofuel production. Here, we have studied the basic mechanisms of cell wall biosynthesis and identified genes involved in biosynthesis of pectic galactan, including the GALS1 galactan synthase andmore » the UDP-galactose/UDP-rhamnose transporter URGT1. We have engineered plants with a more suitable biomass composition by applying these findings, in conjunction with synthetic biology and gene stacking tools. Plants were engineered to have up to fourfold more pectic galactan in stems by overexpressing GALS1, URGT1, and UGE2, a UDP-glucose epimerase. Furthermore, the increased galactan trait was engineered into plants that were already engineered to have low xylan content by restricting xylan biosynthesis to vessels where this polysaccharide is essential. Finally, the high galactan and low xylan traits were stacked with the low lignin trait obtained by expressing the QsuB gene encoding dehydroshikimate dehydratase in lignifying cells. In conclusion, the results show that approaches to increasing C6 sugar content, decreasing xylan, and reducing lignin content can be combined in an additive manner. Thus, the engineered lines obtained by this trait-stacking approach have substantially improved properties from the perspective of biofuel production, and they do not show any obvious negative growth effects. The approach used in this study can be readily transferred to bioenergy crop plants.« less

HOT CELL BUILDING, TRA632, INTERIOR. CONTEXTUAL VIEW OF HOT CELL ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

HOT CELL BUILDING, TRA-632, INTERIOR. CONTEXTUAL VIEW OF HOT CELL NO. 2 FROM STAIRWAY ALONG NORTH WALL. OBSERVATION WINDOW ALONG WEST SIDE BENEATH "CELL 2" SIGN. DOORWAY IN LEFT OF VIEW LEADS TO CELL 1 WORK AREA OR TO EXIT OUTDOORS TO NORTH. RADIATION DETECTION MONITOR TO RIGHT OF DOOR. CAMERA FACING SOUTHWEST. INL NEGATIVE NO. HD46-28-3. Mike Crane, Photographer, 2/2005 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

Fabrication of porous chitosan/poly(vinyl alcohol) reinforced single-walled carbon nanotube nanocomposites for neural tissue engineering.

PubMed

Shokrgozar, Mohammad Ali; Mottaghitalab, Fatemeh; Mottaghitalab, Vahid; Farokhi, Mehdi

2011-04-01

With the ability to form a nano-sized fibrous structure with large pore sizes mimicking the extracellular matrix (ECM), electrospinning was used to fabricate chitosan/poly(vinyl alcohol) nanofibers reinforced by single-walled carbon nanotube (SWNT-CS/PVA) for potential use in neural tissue engineering. Moreover, ultrasonication was performed to fabricate highly dispersed SWNT/CS solution with 7%, 12%, and 17% SWNT content prior to electrospinning process. In the present study, a number of properties of CS/PVA reinforced SWNTs nanocomposites were evaluated. The in vitro biocompatibility of the electrospun fiber mats was also assessed using human brain-derived cells and U373 cell lines. The results have shown that SWNTs as reinforcing phase can augment the morphology, porosity, and structural properties of CS/PVA nanofiber composites and thus benefit the proliferation rate of both cell types. In addition, the cells exhibit their normal morphology while integrating with surrounding fibers. The results confirmed the potential of SWNT-CS/PVA nanocomposites as scaffold for neural tissue engineering.

Genetically Engineered Materials for Biofuels Production

NASA Astrophysics Data System (ADS)

Raab, Michael

2012-02-01

Agrivida, Inc., is an agricultural biotechnology company developing industrial crop feedstocks for the fuel and chemical industries. Agrivida's crops have improved processing traits that enable efficient, low cost conversion of the crops' cellulosic components into fermentable sugars. Currently, pretreatment and enzymatic conversion of the major cell wall components, cellulose and hemicellulose, into fermentable sugars is the most expensive processing step that prevents widespread adoption of biomass in biofuels processes. To lower production costs we are consolidating pretreatment and enzyme production within the crop. In this strategy, transgenic plants express engineered cell wall degrading enzymes in an inactive form, which can be reactivated after harvest. We have engineered protein elements that disrupt enzyme activity during normal plant growth. Upon exposure to specific processing conditions, the engineered enzymes are converted into their active forms. This mechanism significantly lowers pretreatment costs and enzyme loadings (>75% reduction) below those currently available to the industry.

Three-Dimensional Transgenic Cell Models to Quantify Space Genotoxic Effects

NASA Technical Reports Server (NTRS)

Gonda, S.; Wu, H.; Pingerelli, P.; Glickman, B.

2000-01-01

In this paper we describe a three-dimensional, multicellular tissue-equivalent model, produced in NASA-designed, rotating wall bioreactors using mammalian cells engineered for genomic containment of mUltiple copies of defined target genes for genotoxic assessment. The Rat 2(lambda) fibroblasts (Stratagene, Inc.) were genetically engineered to contain high-density target genes for mutagenesis. Stable three-dimensional, multicellular spheroids were formed when human mammary epithelial cells and Rat 2(lambda) fibroblasts were cocultured on Cytodex 3 Beads in a rotating wall bioreactor. The utility of this spheroidal model for genotoxic assessment was indicated by a linear dose response curve and by results of gene sequence analysis of mutant clones from 400micron diameter spheroids following low-dose, high-energy, neon radiation exposure

Magnetically actuated tissue engineered scaffold: insights into mechanism of physical stimulation

NASA Astrophysics Data System (ADS)

Sapir-Lekhovitser, Yulia; Rotenberg, Menahem Y.; Jopp, Juergen; Friedman, Gary; Polyak, Boris; Cohen, Smadar

2016-02-01

Providing the right stimulatory conditions resulting in efficient tissue promoting microenvironment in vitro and in vivo is one of the ultimate goals in tissue development for regenerative medicine. It has been shown that in addition to molecular signals (e.g. growth factors) physical cues are also required for generation of functional cell constructs. These cues are particularly relevant to engineering of biological tissues, within which mechanical stress activates mechano-sensitive receptors, initiating biochemical pathways which lead to the production of functionally mature tissue. Uniform magnetic fields coupled with magnetizable nanoparticles embedded within three dimensional (3D) scaffold structures remotely create transient physical forces that can be transferrable to cells present in close proximity to the nanoparticles. This study investigated the hypothesis that magnetically responsive alginate scaffold can undergo reversible shape deformation due to alignment of scaffold's walls in a uniform magnetic field. Using custom made Helmholtz coil setup adapted to an Atomic Force Microscope we monitored changes in matrix dimensions in situ as a function of applied magnetic field, concentration of magnetic particles within the scaffold wall structure and rigidity of the matrix. Our results show that magnetically responsive scaffolds exposed to an externally applied time-varying uniform magnetic field undergo a reversible shape deformation. This indicates on possibility of generating bending/stretching forces that may exert a mechanical effect on cells due to alternating pattern of scaffold wall alignment and relaxation. We suggest that the matrix structure deformation is produced by immobilized magnetic nanoparticles within the matrix walls resulting in a collective alignment of scaffold walls upon magnetization. The estimated mechanical force that can be imparted on cells grown on the scaffold wall at experimental conditions is in the order of 1 pN, which correlates well with reported threshold to induce mechanotransduction effects on cellular level. This work is our next step in understanding of how to accurately create proper stimulatory microenvironment for promotion of cellular organization to form mature tissue engineered constructs.

Magnetically actuated tissue engineered scaffold: insights into mechanism of physical stimulation

PubMed Central

Sapir-Lekhovitser, Yulia; Rotenberg, Menahem Y.; Jopp, Juergen; Friedman, Gary; Polyak, Boris; Cohen, Smadar

2016-01-01

Providing the right stimulatory conditions resulting in efficient tissue promoting microenvironment in vitro and in vivo is one of the ultimate goals in tissue development for regenerative medicine. It has been shown that in addition to molecular signals (e.g. growth factors) physical cues are also required for generation of functional cell constructs. These cues are particularly relevant to engineering of biological tissues, within which mechanical stress activates mechano-sensitive receptors, initiating biochemical pathways which lead to the production of functionally mature tissue. Uniform magnetic fields coupled with magnetizable nanoparticles embedded within three dimensional (3D) scaffold structures remotely create transient physical forces that can be transferrable to cells present in close proximity to the nanoparticles. This study investigated the hypothesis that magnetically responsive alginate scaffold can undergo reversible shape deformation due to alignment of scaffold’s walls in a uniform magnetic field. Using custom made Helmholtz coil setup adapted to an Atomic Force Microscope we monitored changes in matrix dimensions in situ as a function of applied magnetic field, concentration of magnetic particles within the scaffold wall structure and rigidity of the matrix. Our results show that magnetically responsive scaffolds exposed to an externally applied time-varying uniform magnetic field undergo a reversible shape deformation. This indicates on possibility of generating bending/stretching forces that may exert a mechanical effect on cells due to alternating pattern of scaffold wall alignment and relaxation. We suggest that the matrix structure deformation is produced by immobilized magnetic nanoparticles within the matrix walls resulting in a collective alignment of scaffold walls upon magnetization. The estimated mechanical force that can be imparted on cells grown on the scaffold wall at experimental conditions is in the order of 1 pN, which correlates well with reported threshold to induce mechanotransduction effects on cellular level. This work is our next step in understanding of how to accurately create proper stimulatory microenvironment for promotion of cellular organization to form mature tissue engineered constructs. PMID:26790538

Genetic Engineering of Maize (Zea mays L.) with Improved Grain Nutrients.

PubMed

Guo, Xiaotong; Duan, Xiaoguang; Wu, Yongzhen; Cheng, Jieshan; Zhang, Juan; Zhang, Hongxia; Li, Bei

2018-02-21

Cell-wall invertase plays important roles in the grain filling of crop plants. However, its functions in the improvement of grain nutrients have not been investigated. In this work, the stable expression of cell-wall-invertase-encoding genes from different plant species and the contents of total starch, protein, amino acid, nitrogen, lipid, and phosphorus were examined in transgenic maize plants. High expressions of the cell-wall-invertase gene conferred enhanced invertase activity and sugar content in transgenic plants, leading to increased grain yield and improved grain nutrients. Transgenic plants with high expressions of the transgene produced more total starch, protein, nitrogen, and essential amino acids in the seeds. Overall, the results indicate that the cell-wall-invertase gene can be used as a potential candidate for the genetic breeding of grain crops with both improved grain yield and quality.

CONSTRUCTION PROGRESS PHOTO OF HOT PILOT PLANT (CP640) LOOKING NORTHWEST ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

CONSTRUCTION PROGRESS PHOTO OF HOT PILOT PLANT (CP-640) LOOKING NORTHWEST SHOWING FORMING AND PLACEMENT OF REINFORCING STEEL FOR SOUTH WALLS OF CELLS 1, 3, 4 AND 5 AND WEST WALL FOR CELLS 1 AND 2; CONSTRUCTION 13 PERCENT COMPLETE. INL PHOTO NUMBER NRTS 59-6436. J. Anderson, Photographer, 12/18/1959 - Idaho National Engineering Laboratory, Idaho Chemical Processing Plant, Fuel Reprocessing Complex, Scoville, Butte County, ID

13C cell wall enrichment and ionic liquid NMR analysis: progress towards a high-throughput detailed chemical analysis of the whole plant cell wall.

PubMed

Foston, Marcus; Samuel, Reichel; Ragauskas, Arthur J

2012-09-07

The ability to accurately and rapidly measure plant cell wall composition, relative monolignol content and lignin-hemicellulose inter-unit linkage distributions has become essential to efforts centered on reducing the recalcitrance of biomass by genetic engineering. Growing (13)C enriched transgenic plants is a viable route to achieve the high-throughput, detailed chemical analysis of whole plant cell wall before and after pretreatment and microbial or enzymatic utilization by (13)C nuclear magnetic resonance (NMR) in a perdeuterated ionic liquid solvent system not requiring component isolation. 1D (13)C whole cell wall ionic liquid NMR of natural abundant and (13)C enriched corn stover stem samples suggest that a high level of uniform labeling (>97%) can significantly reduce the total NMR experiment times up to ~220 times. Similarly, significant reduction in total NMR experiment time (~39 times) of the (13)C enriched corn stover stem samples for 2D (13)C-(1)H heteronuclear single quantum coherence NMR was found.

A versatile strategy for grafting polymers to wood cell walls.

PubMed

Keplinger, T; Cabane, E; Chanana, M; Hass, P; Merk, V; Gierlinger, N; Burgert, I

2015-01-01

The hierarchical structure of wood is composed of a cellulose skeleton of high structural order at various length scales. At the nanoscale and microscale the specific structural features of the cells and cell walls result in a lightweight structure with an anisotropic material profile of excellent mechanical performance. By being able to specifically functionalize wood at the level of cell and cell walls one can insert new properties and inevitably upscale them along the intrinsic hierarchical structure, to a level of large-scale engineering materials applications. For this purpose, however, precise control of the spatial distribution of the modifying substances in the complex wood structure is needed. Here we demonstrate a method to insert methacryl groups into wood cell walls using two different chemistry routes. By using these methacryl groups as the anchor points for grafting, various polymers can be inserted into the wood structure. Strikingly, depending on the methacryl precursor, the spatial distribution of the polymer differs strongly. As a proof of concept we grafted polystyrene as a model compound in the second modification step. In the case of methacryloyl chloride the polymer was located mainly at the interface between the cell lumina and the cell wall covering the inner surface of the cells and being traceable up to 2-3 μm in the cell wall, whereas in the case of methacrylic anhydride the polymer was located inside the whole cell wall. Scanning electron microscopy, Fourier transform infrared spectroscopy and especially Raman spectroscopy were used for an in-depth analysis of the modified wood at the cell wall level. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

LPT. Low power test control building (TAN641) interior. Camera facing ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

LPT. Low power test control building (TAN-641) interior. Camera facing northeast at what remains of control room console. Cut in wall at right of view shows west wall of northern test cell. INEEL negative no. HD-40-4-4 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gaul, Alexander; Holzinger, Dennis; Müglich, Nicolas David

A magnetic domain texture has been deterministically engineered in a topographically flat exchange-biased (EB) thin film system. The texture consists of long-range periodically arranged unit cells of four individual domains, characterized by individual anisotropies, individual geometry, and with non-collinear remanent magnetizations. The texture has been engineered by a sequence of light-ion bombardment induced magnetic patterning of the EB layer system. The magnetic texture's in-plane spatial magnetization distribution and the corresponding domain walls have been characterized by scanning electron microscopy with polarization analysis (SEMPA). The influence of magnetic stray fields emerging from neighboring domain walls and the influence of the differentmore » anisotropies of the adjacent domains on the Néel type domain wall core's magnetization rotation sense and widths were investigated. It is shown that the usual energy degeneracy of clockwise and counterclockwise rotating magnetization through the walls is revoked, suppressing Bloch lines along the domain wall. Estimates of the domain wall widths for different domain configurations based on material parameters determined by vibrating sample magnetometry were quantitatively compared to the SEMPA data.« less

Systems and synthetic biology approaches to alter plant cell walls and reduce biomass recalcitrance

DOE PAGES

Kalluri, Udaya C.; Yin, Hengfu; Yang, Xiaohan; ...

2014-11-03

Fine-tuning plant cell wall properties to render plant biomass more amenable to biofuel conversion is a colossal challenge. A deep knowledge of the biosynthesis and regulation of plant cell wall and a high-precision genome engineering toolset are the two essential pillars of efforts to alter plant cell walls and reduce biomass recalcitrance. The past decade has seen a meteoric rise in use of transcriptomics and high-resolution imaging methods resulting in fresh insights into composition, structure, formation and deconstruction of plant cell walls. Subsequent gene manipulation approaches, however, commonly include ubiquitous mis-expression of a single candidate gene in a host thatmore » carries an intact copy of the native gene. The challenges posed by pleiotropic and unintended changes resulting from such an approach are moving the field towards synthetic biology approaches. Finally, synthetic biology builds on a systems biology knowledge base and leverages high-precision tools for high-throughput assembly of multigene constructs and pathways, precision genome editing and site-specific gene stacking, silencing and/or removal. Here, we summarize the recent breakthroughs in biosynthesis and remodelling of major secondary cell wall components, assess the impediments in obtaining a systems-level understanding and explore the potential opportunities in leveraging synthetic biology approaches to reduce biomass recalcitrance.« less

Systems and synthetic biology approaches to alter plant cell walls and reduce biomass recalcitrance

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kalluri, Udaya C.; Yin, Hengfu; Yang, Xiaohan

Fine-tuning plant cell wall properties to render plant biomass more amenable to biofuel conversion is a colossal challenge. A deep knowledge of the biosynthesis and regulation of plant cell wall and a high-precision genome engineering toolset are the two essential pillars of efforts to alter plant cell walls and reduce biomass recalcitrance. The past decade has seen a meteoric rise in use of transcriptomics and high-resolution imaging methods resulting in fresh insights into composition, structure, formation and deconstruction of plant cell walls. Subsequent gene manipulation approaches, however, commonly include ubiquitous mis-expression of a single candidate gene in a host thatmore » carries an intact copy of the native gene. The challenges posed by pleiotropic and unintended changes resulting from such an approach are moving the field towards synthetic biology approaches. Finally, synthetic biology builds on a systems biology knowledge base and leverages high-precision tools for high-throughput assembly of multigene constructs and pathways, precision genome editing and site-specific gene stacking, silencing and/or removal. Here, we summarize the recent breakthroughs in biosynthesis and remodelling of major secondary cell wall components, assess the impediments in obtaining a systems-level understanding and explore the potential opportunities in leveraging synthetic biology approaches to reduce biomass recalcitrance.« less

Enzymatic cell wall degradation of Chlorella vulgaris and other microalgae for biofuels production.

PubMed

Gerken, Henri G; Donohoe, Bryon; Knoshaug, Eric P

2013-01-01

Cell walls of microalgae consist of a polysaccharide and glycoprotein matrix providing the cells with a formidable defense against its environment. We characterized enzymes that can digest the cell wall and weaken this defense for the purpose of protoplasting or lipid extraction. A growth inhibition screen demonstrated that chitinase, lysozyme, pectinase, sulfatase, β-glucuronidase, and laminarinase had the broadest effect across the various Chlorella strains tested and also inhibited Nannochloropsis and Nannochloris strains. Chlorella is typically most sensitive to chitinases and lysozymes, both enzymes that degrade polymers containing N-acetylglucosamine. Using a fluorescent DNA stain, we developed rapid methodology to quantify changes in permeability in response to enzyme digestion and found that treatment with lysozyme in conjunction with other enzymes has a drastic effect on cell permeability. Transmission electron microscopy of enzymatically treated Chlorella vulgaris indicates that lysozyme degrades the outer surface of the cell wall and removes hair-like fibers protruding from the surface, which differs from the activity of chitinase. This action on the outer surface of the cell causes visible protuberances on the cell surface and presumably leads to the increased settling rate when cells are treated with lysozyme. We demonstrate radical ultrastructural changes to the cell wall in response to treatment with various enzyme combinations which, in some cases, causes a greater than twofold increase in the thickness of the cell wall. The enzymes characterized in this study should prove useful in the engineering and extraction of oils from microalgae.

In vitro evaluation of three-dimensional single-walled carbon nanotube composites for bone tissue engineering.

PubMed

Gupta, Ashim; Main, Benjamin J; Taylor, Brittany L; Gupta, Manu; Whitworth, Craig A; Cady, Craig; Freeman, Joseph W; El-Amin, Saadiq F

2014-11-01

The purpose of this study was to develop three-dimensional single-walled carbon nanotube composites (SWCNT/PLAGA) using 10-mg single-walled carbon nanotubes (SWCNT) for bone regeneration and to determine the mechanical strength of the composites, and to evaluate the interaction of MC3T3-E1 cells via cell adhesion, growth, survival, proliferation, and gene expression. PLAGA (polylactic-co-glycolic acid) and SWCNT/PLAGA microspheres and composites were fabricated, characterized, and mechanical testing was performed. MC3T3-E1 cells were seeded and cell adhesion/morphology, growth/survival, proliferation, and gene expression analysis were performed to evaluate biocompatibility. Imaging studies demonstrated microspheres with uniform shape and smooth surfaces, and uniform incorporation of SWCNT into PLAGA matrix. The microspheres bonded in a random packing manner while maintaining spacing, thus resembling trabeculae of cancellous bone. Addition of SWCNT led to greater compressive modulus and ultimate compressive strength. Imaging studies revealed that MC3T3-E1 cells adhered, grew/survived, and exhibited normal, nonstressed morphology on the composites. SWCNT/PLAGA composites exhibited higher cell proliferation rate and gene expression compared with PLAGA. These results demonstrate the potential of SWCNT/PLAGA composites for musculoskeletal regeneration, for bone tissue engineering, and are promising for orthopedic applications as they possess the combined effect of increased mechanical strength, cell proliferation, and gene expression. © 2014 Wiley Periodicals, Inc.

Deletion of Aspergillus nidulans GDP-mannose transporters affects hyphal morphometry, cell wall architecture, spore surface character, cell adhesion, and biofilm formation.

PubMed

Kadry, Ashraf A; El-Ganiny, Amira M; Mosbah, Rasha A; Kaminskyj, Susan G W

2018-07-01

Systemic human fungal infections are increasingly common. Aspergillus species cause most of the airborne fungal infections. Life-threatening invasive aspergillosis was formerly found only in immune-suppressed patients, but recently some strains of A. fumigatus have become primary pathogens. Many fungal cell wall components are absent from mammalian systems, so they are potential drug targets. Cell-wall-targeting drugs such as echinocandins are used clinically, although echinocandin-resistant strains were discovered shortly after their introduction. Currently there are no fully effective anti-fungal drugs. Fungal cell wall glycoconjugates modulate human immune responses, as well as fungal cell adhesion, biofilm formation, and drug resistance. Guanosine diphosphate (GDP) mannose transporters (GMTs) transfer GDP-mannose from the cytosol to the Golgi lumen prior to mannosylation. Aspergillus nidulans GMTs are encoded by gmtA and gmtB. Here we elucidate the roles of A. nidulans GMTs. Strains engineered to lack either or both GMTs were assessed for hyphal and colonial morphology, cell wall ultrastructure, antifungal susceptibility, spore hydrophobicity, adherence and biofilm formation. The gmt-deleted strains had smaller colonies with reduced sporulation and with thicker hyphal walls. The gmtA deficient spores had reduced hydrophobicity and were less adherent and less able to form biofilms in vitro. Thus, gmtA not only participates in maintaining the cell wall integrity but also plays an important role in biofilm establishment and adherence of A. nidulans. These findings suggested that GMTs have roles in A. nidulans growth and cell-cell interaction and could be a potential target for new antifungals that target virulence determinants.

Plant cell wall sugars: sweeteners for a bio-based economy.

PubMed

Van de Wouwer, Dorien; Boerjan, Wout; Vanholme, Bartel

2018-02-12

Global warming and the consequent climate change is one of the major environmental challenges we are facing today. The driving force behind the rise in temperature is our fossil-based economy, which releases massive amounts of the greenhouse gas carbon dioxide into the atmosphere. In order to reduce greenhouse gas emission, we need to scale down our dependency on fossil resources, implying that we need other sources for energy and chemicals to feed our economy. Here, plants have an important role to play; by means of photosynthesis, plants capture solar energy to split water and fix carbon derived from atmospheric carbon dioxide. A significant fraction of the fixed carbon ends up as polysaccharides in the plant cell wall. Fermentable sugars derived from cell wall polysaccharides form an ideal carbon source for the production of bio-platform molecules. However, a major limiting factor in the use of plant biomass as feedstock for the bio-based economy is the complexity of the plant cell wall and its recalcitrance towards deconstruction. To facilitate the release of fermentable sugars during downstream biomass processing, the composition and structure of the cell wall can be engineered. Different strategies to reduce cell wall recalcitrance will be described in this review. The ultimate goal is to obtain a tailor-made biomass, derived from plants with a cell wall optimized for particular industrial or agricultural applications, without affecting plant growth and development. This article is protected by copyright. All rights reserved.

Target or barrier? The cell wall of early- and later-diverging plants vs cadmium toxicity: differences in the response mechanisms

PubMed Central

Parrotta, Luigi; Guerriero, Gea; Sergeant, Kjell; Cai, Giampiero; Hausman, Jean-Francois

2015-01-01

Increasing industrialization and urbanization result in emission of pollutants in the environment including toxic heavy metals, as cadmium and lead. Among the different heavy metals contaminating the environment, cadmium raises great concern, as it is ecotoxic and as such can heavily impact ecosystems. The cell wall is the first structure of plant cells to come in contact with heavy metals. Its composition, characterized by proteins, polysaccharides and in some instances lignin and other phenolic compounds, confers the ability to bind non-covalently and/or covalently heavy metals via functional groups. A strong body of evidence in the literature has shown the role of the cell wall in heavy metal response: it sequesters heavy metals, but at the same time its synthesis and composition can be severely affected. The present review analyzes the dual property of plant cell walls, i.e., barrier and target of heavy metals, by taking Cd toxicity as example. Following a summary of the known physiological and biochemical responses of plants to Cd, the review compares the wall-related mechanisms in early- and later-diverging land plants, by considering the diversity in cell wall composition. By doing so, common as well as unique response mechanisms to metal/cadmium toxicity are identified among plant phyla and discussed. After discussing the role of hyperaccumulators’ cell walls as a particular case, the review concludes by considering important aspects for plant engineering. PMID:25814996

13. VIEW OF EAST OPERATING GALLERY ALONG THE POSTMORTEM CELLS. ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

13. VIEW OF EAST OPERATING GALLERY ALONG THE POST-MORTEM CELLS. A NUMBER OF MANIPULATOR ARMS COVERED WITH PLASTIC ARE ON THE LEFT WALL. - Nevada Test Site, Engine Maintenance Assembly & Disassembly Facility, Area 25, Jackass Flats, Mercury, Nye County, NV

Adding dimension to cellular mechanotransduction: Advances in biomedical engineering of multiaxial cell-stretch systems and their application to cardiovascular biomechanics and mechano-signaling.

PubMed

Friedrich, O; Schneidereit, D; Nikolaev, Y A; Nikolova-Krstevski, V; Schürmann, S; Wirth-Hücking, A; Merten, A L; Fatkin, D; Martinac, B

2017-11-01

Hollow organs (e.g. heart) experience pressure-induced mechanical wall stress sensed by molecular mechano-biosensors, including mechanosensitive ion channels, to translate into intracellular signaling. For direct mechanistic studies, stretch devices to apply defined extensions to cells adhered to elastomeric membranes have stimulated mechanotransduction research. However, most engineered systems only exploit unilateral cellular stretch. In addition, it is often taken for granted that stretch applied by hardware translates 1:1 to the cell membrane. However, the latter crucially depends on the tightness of the cell-substrate junction by focal adhesion complexes and is often not calibrated for. In the heart, (increased) hemodynamic volume/pressure load is associated with (increased) multiaxial wall tension, stretching individual cardiomyocytes in multiple directions. To adequately study cellular models of chronic organ distension on a cellular level, biomedical engineering faces challenges to implement multiaxial cell stretch systems that allow observing cell reactions to stretch during live-cell imaging, and to calibrate for hardware-to-cell membrane stretch translation. Here, we review mechanotransduction, cell stretch technologies from uni-to multiaxial designs in cardio-vascular research, and the importance of the stretch substrate-cell membrane junction. We also present new results using our IsoStretcher to demonstrate mechanosensitivity of Piezo1 in HEK293 cells and stretch-induced Ca 2+ entry in 3D-hydrogel-embedded cardiomyocytes. Copyright © 2017 Elsevier Ltd. All rights reserved.

Constitutive expression of cell wall invertase genes increases grain yield and starch content in maize.

PubMed

Li, Bei; Liu, Hua; Zhang, Yue; Kang, Tao; Zhang, Li; Tong, Jianhua; Xiao, Langtao; Zhang, Hongxia

2013-12-01

Grain size, number and starch content are important determinants of grain yield and quality. One of the most important biological processes that determine these components is the carbon partitioning during the early grain filling, which requires the function of cell wall invertase. Here, we showed the constitutive expression of cell wall invertase-encoding gene from Arabidopsis, rice (Oryza sativa) or maize (Zea mays), driven by the cauliflower mosaic virus (CaMV) 35S promoter, all increased cell wall invertase activities in different tissues and organs, including leaves and developing seeds, and substantially improved grain yield up to 145.3% in transgenic maize plants as compared to the wild-type plants, an effect that was reproduced in our 2-year field trials at different locations. The dramatically increased grain yield is due to the enlarged ears with both enhanced grain size and grain number. Constitutive expression of the invertase-encoding gene also increased total starch content up to 20% in the transgenic kernels. Our results suggest that cell wall invertase gene can be genetically engineered to improve both grain yield and grain quality in crop plants. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Chimeric Antigen Receptor- and TCR-Modified T Cells Enter Main Street and Wall Street.

PubMed

Barrett, David M; Grupp, Stephan A; June, Carl H

2015-08-01

The field of adoptive cell transfer (ACT) is currently comprised of chimeric Ag receptor (CAR)- and TCR-engineered T cells and has emerged from principles of basic immunology to paradigm-shifting clinical immunotherapy. ACT of T cells engineered to express artificial receptors that target cells of choice is an exciting new approach for cancer, and it holds equal promise for chronic infection and autoimmunity. Using principles of synthetic biology, advances in immunology, and genetic engineering have made it possible to generate human T cells that display desired specificities and enhanced functionalities. Clinical trials in patients with advanced B cell leukemias and lymphomas treated with CD19-specific CAR T cells have induced durable remissions in adults and children. The prospects for the widespread availability of engineered T cells have changed dramatically given the recent entry of the pharmaceutical industry to this arena. In this overview, we discuss some of the challenges and opportunities that face the field of ACT. Copyright © 2015 by The American Association of Immunologists, Inc.

Colorectal tissue engineering: A comparative study between porcine small intestinal submucosa (SIS) and chitosan hydrogel patches.

PubMed

Denost, Quentin; Adam, Jean-Philippe; Pontallier, Arnaud; Montembault, Alexandra; Bareille, Reine; Siadous, Robin; Delmond, Samantha; Rullier, Eric; David, Laurent; Bordenave, Laurence

2015-12-01

Tissue engineering may provide new operative tools for colorectal surgery in elective indications. The aim of this study was to define a suitable bioscaffold for colorectal tissue engineering. We compared 2 bioscaffolds with in vitro and in vivo experiments: porcine small intestinal submucosa (SIS) versus chitosan hydrogel matrix. We assessed nontoxicity of the scaffold in vitro by using human adipose-derived stem cells (hADSC). In vivo, a 1 × 2-cm colonic wall defect was created in 16 rabbits. Animals were divided randomly into 2 groups according to the graft used, SIS or chitosan hydrogel. Graft area was explanted at 4 and 8 weeks. The end points of in vivo experiments were technical feasibility, behavior of the scaffold, in situ putative inflammatory effect, and the quality of tissue regeneration, in particular smooth muscle layer regeneration. In vitro, hADSC attachment and proliferation occurred on both scaffolds without a substantial difference. After proliferation, hADSCs kept their mesenchymal stem cell characteristics. In vivo, one animal died in each group. Eight weeks after implantation, the chitosan scaffold allowed better wound healing compared with the SIS scaffold, with more effective control of inflammatory activity and an integral regeneration of the colonic wall including the smooth muscle cell layer. The outcomes of in vitro experiments did not differ greatly between the 2 groups. Macroscopic and histologic findings, however, revealed better wound healing of the colonic wall in the chitosan group suggesting that the chitosan hydrogel could serve as a better scaffold for colorectal tissue engineering. Copyright © 2015 Elsevier Inc. All rights reserved.

Biomineralization of gold by Mucor plumbeus: The progress in understanding the mechanism of nanoparticles' formation.

PubMed

Maliszewska, Irena; Tylus, Włodzimierz; Chęcmanowski, Jacek; Szczygieł, Bogdan; Pawlaczyk-Graja, Izabela; Pusz, Wojciech; Baturo-Cieśniewska, Anna

2017-09-01

This contribution describes the deposition of gold nanoparticles by microbial reduction of Au(III) ions using the mycelium of Mucor plumbeus. Biosorption as the major mechanism of Au(III) ions binding by the fungal cells and the reduction of them to the form of Au(0) on/in the cell wall, followed by the transportation of the synthesized gold nanoparticles to the cytoplasm, is postulated. The probable mechanism behind the reduction of Au(III) ions is discussed, leading to the conclusion that this process is nonenzymatic one. Chitosan of the fungal cell wall is most likely to be the major molecule involved in biomineralization of gold by the mycelium of M. plumbeus. Separation of gold nanoparticles from the cells has been carried out by the ultrasonic disintegration and the obtained nanostructures were characterized by UV-vis spectroscopy and transmission electron micrograph analysis. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1381-1392, 2017. © 2017 American Institute of Chemical Engineers.

Colony Organization in the Green Alga Botryococcus braunii (Race B) Is Specified by a Complex Extracellular Matrix

PubMed Central

Weiss, Taylor L.; Roth, Robyn; Goodson, Carrie; Vitha, Stanislav; Black, Ian; Azadi, Parastoo; Rusch, Jannette; Holzenburg, Andreas

2012-01-01

Botryococcus braunii is a colonial green alga whose cells associate via a complex extracellular matrix (ECM) and produce prodigious amounts of liquid hydrocarbons that can be readily converted into conventional combustion engine fuels. We used quick-freeze deep-etch electron microscopy and biochemical/histochemical analysis to elucidate many new features of B. braunii cell/colony organization and composition. Intracellular lipid bodies associate with the chloroplast and endoplasmic reticulum (ER) but show no evidence of being secreted. The ER displays striking fenestrations and forms a continuous subcortical system in direct contact with the cell membrane. The ECM has three distinct components. (i) Each cell is surrounded by a fibrous β-1, 4- and/or β-1, 3-glucan-containing cell wall. (ii) The intracolonial ECM space is filled with a cross-linked hydrocarbon network permeated with liquid hydrocarbons. (iii) Colonies are enclosed in a retaining wall festooned with a fibrillar sheath dominated by arabinose-galactose polysaccharides, which sequesters ECM liquid hydrocarbons. Each cell apex associates with the retaining wall and contributes to its synthesis. Retaining-wall domains also form “drapes” between cells, with some folding in on themselves and penetrating the hydrocarbon interior of a mother colony, partitioning it into daughter colonies. We propose that retaining-wall components are synthesized in the apical Golgi apparatus, delivered to apical ER fenestrations, and assembled on the surfaces of apical cell walls, where a proteinaceous granular layer apparently participates in fibril morphogenesis. We further propose that hydrocarbons are produced by the nonapical ER, directly delivered to the contiguous cell membrane, and pass across the nonapical cell wall into the hydrocarbon-based ECM. PMID:22941913

Colony organization in the green alga Botryococcus braunii is specified by a complex extracellular matrix

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weiss, Taylor L.; Roth, Robyn; Goodson, Carrie

Botryococcus braunii is a colonial green alga whose cells associate via a complex extracellular matrix (ECM) and produce prodigious amounts of liquid hydrocarbons that can be readily converted into conventional combustion engine fuels. We used quickfreeze deep-etch electron microscopy and biochemical/histochemical analysis to elucidate many new features of B. braunii cell/colony organization and composition. Intracellular lipid bodies associate with the chloroplast and endoplasmic reticulum (ER) but show no evidence of being secreted. The ER displays striking fenestrations and forms a continuous subcortical system in direct contact with the cell membrane. The ECM has three distinct components. (i) Each cell ismore » surrounded by a fibrous β-1, 4- and/or β-1, 3-glucan-containing cell wall. (ii) The intracolonial ECM space is filled with a cross-linked hydrocarbon network permeated with liquid hydrocarbons. (iii) Colonies are enclosed in a retaining wall festooned with a fibrillar sheath dominated by arabinose-galactose polysaccharides, which sequesters ECM liquid hydrocarbons. Each cell apex associates with the retaining wall and contributes to its synthesis. Retaining-wall domains also form "drapes" between cells, with some folding in on themselves and penetrating the hydrocarbon interior of a mother colony, partitioning it into daughter colonies. In addition, we propose that retaining-wall components are synthesized in the apical Golgi apparatus, delivered to apical ER fenestrations, and assembled on the surfaces of apical cell walls, where a proteinaceous granular layer apparently participates in fibril morphogenesis. We further propose that hydrocarbons are produced by the nonapical ER, directly delivered to the contiguous cell membrane, and pass across the nonapical cell wall into the hydrocarbon-based ECM.« less

Physiological and Transcriptional Responses of Saccharomyces cerevisiae to d-Limonene Show Changes to the Cell Wall but Not to the Plasma Membrane

PubMed Central

Brennan, Timothy C. R.; Nielsen, Lars K.

2013-01-01

Monoterpenes can, upon hydrogenation, be used as light-fraction components of sustainable aviation fuels. Fermentative production of monoterpenes in engineered microorganisms, such as Saccharomyces cerevisiae, has gained attention as a potential route to deliver these next-generation fuels from renewable biomass. However, end product toxicity presents a formidable problem for microbial synthesis. Due to their hydrophobicity, monoterpene inhibition has long been attributed to membrane interference, but the molecular mechanism remains largely unsolved. In order to gain a better understanding of the mode of action, we analyzed the composition and structural integrity of the cell envelope as well as the transcriptional response of yeast cells treated with an inhibitory amount of d-limonene (107 mg/liter). We found no alterations in membrane fluidity, structural membrane integrity, or fatty acid composition after the solvent challenge. A 4-fold increase in the mean fluorescence intensity per cell (using calcofluor white stain) and increased sensitivity to cell wall-degrading enzymes demonstrated that limonene disrupts cell wall properties. Global transcript measurements confirmed the membrane integrity observations by showing no upregulation of ergosterol or fatty acid biosynthesis pathways, which are commonly overexpressed in yeast to reinforce membrane rigidity during ethanol exposure. Limonene shock did cause a compensatory response to cell wall damage through overexpression of several genes (ROM1, RLM1, PIR3, CTT1, YGP1, MLP1, PST1, and CWP1) involved with the cell wall integrity signaling pathway. This is the first report demonstrating that cell wall, rather than plasma membrane, deterioration is the main source of monoterpene inhibition. We show that limonene can alter the structure and function of the cell wall, which has a clear effect on cytokinesis. PMID:23542628

Colony organization in the green alga Botryococcus braunii is specified by a complex extracellular matrix

DOE PAGES

Weiss, Taylor L.; Roth, Robyn; Goodson, Carrie; ...

2012-08-31

Botryococcus braunii is a colonial green alga whose cells associate via a complex extracellular matrix (ECM) and produce prodigious amounts of liquid hydrocarbons that can be readily converted into conventional combustion engine fuels. We used quickfreeze deep-etch electron microscopy and biochemical/histochemical analysis to elucidate many new features of B. braunii cell/colony organization and composition. Intracellular lipid bodies associate with the chloroplast and endoplasmic reticulum (ER) but show no evidence of being secreted. The ER displays striking fenestrations and forms a continuous subcortical system in direct contact with the cell membrane. The ECM has three distinct components. (i) Each cell ismore » surrounded by a fibrous β-1, 4- and/or β-1, 3-glucan-containing cell wall. (ii) The intracolonial ECM space is filled with a cross-linked hydrocarbon network permeated with liquid hydrocarbons. (iii) Colonies are enclosed in a retaining wall festooned with a fibrillar sheath dominated by arabinose-galactose polysaccharides, which sequesters ECM liquid hydrocarbons. Each cell apex associates with the retaining wall and contributes to its synthesis. Retaining-wall domains also form "drapes" between cells, with some folding in on themselves and penetrating the hydrocarbon interior of a mother colony, partitioning it into daughter colonies. In addition, we propose that retaining-wall components are synthesized in the apical Golgi apparatus, delivered to apical ER fenestrations, and assembled on the surfaces of apical cell walls, where a proteinaceous granular layer apparently participates in fibril morphogenesis. We further propose that hydrocarbons are produced by the nonapical ER, directly delivered to the contiguous cell membrane, and pass across the nonapical cell wall into the hydrocarbon-based ECM.« less

Fabrication of chitin-chitosan/nano TiO2-composite scaffolds for tissue engineering applications.

PubMed

Jayakumar, R; Ramachandran, Roshni; Divyarani, V V; Chennazhi, K P; Tamura, H; Nair, S V

2011-03-01

In this study, we prepared chitin-chitosan/nano TiO(2) composite scaffolds using lyophilization technique for bone tissue engineering. The prepared composite scaffold was characterized using SEM, XRD, FTIR and TGA. In addition, swelling, degradation and biomineralization capability of the composite scaffolds were evaluated. The developed composite scaffold showed controlled swelling and degradation when compared to the control scaffold. Cytocompatibility of the scaffold was assessed by MTT assay and cell attachment studies using osteoblast-like cells (MG-63), fibroblast cells (L929) and human mesenchymal stem cells (hMSCs). Results indicated no sign of toxicity and cells were found attached to the pore walls within the scaffolds. These results suggested that the developed composite scaffold possess the prerequisites for tissue engineering scaffolds and it can be used for tissue engineering applications. Copyright © 2010 Elsevier B.V. All rights reserved.

Wood formation in Angiosperms.

PubMed

Déjardin, Annabelle; Laurans, Françoise; Arnaud, Dominique; Breton, Christian; Pilate, Gilles; Leplé, Jean-Charles

2010-04-01

Wood formation is a complex biological process, involving five major developmental steps, including (1) cell division from a secondary meristem called the vascular cambium, (2) cell expansion (cell elongation and radial enlargement), (3) secondary cell wall deposition, (4) programmed cell death, and (5) heartwood formation. Thanks to the development of genomic studies in woody species, as well as genetic engineering, recent progress has been made in the understanding of the molecular mechanisms underlying wood formation. In this review, we will focus on two different aspects, the lignification process and the control of microfibril angle in the cell wall of wood fibres, as they are both key features of wood material properties. Copyright (c) 2010 Académie des sciences. Published by Elsevier SAS. All rights reserved.

PCR on yeast colonies: an improved method for glyco-engineered Saccharomyces cerevisiae

PubMed Central

2013-01-01

Background Saccharomyces cerevisiae is extensively used in bio-industries. However, its genetic engineering to introduce new metabolism pathways can cause unexpected phenotypic alterations. For example, humanisation of the glycosylation pathways is a high priority pharmaceutical industry goal for production of therapeutic glycoproteins in yeast. Genomic modifications can lead to several described physiological changes: biomass yields decrease, temperature sensitivity or cell wall structure modifications. We have observed that deletion of several N-mannosyltransferases in Saccharomyces cerevisiae, results in strains that can no longer be analyzed by classical PCR on yeast colonies. Findings In order to validate our glyco-engineered Saccharomyces cerevisiae strains, we developed a new protocol to carry out PCR directly on genetically modified yeast colonies. A liquid culture phase, combined with the use of a Hot Start DNA polymerase, allows a 3-fold improvement of PCR efficiency. The results obtained are repeatable and independent of the targeted sequence; as such the protocol is well adapted for intensive screening applications. Conclusions The developed protocol enables by-passing of many of the difficulties associated with PCR caused by phenotypic modifications brought about by humanisation of the glycosylation in yeast and allows rapid validation of glyco-engineered Saccharomyces cerevisiae cells. It has the potential to be extended to other yeast strains presenting cell wall structure modifications. PMID:23688076

Antibiotic Algae by Chemical Surface Engineering.

PubMed

Kerschgens, Isabel P; Gademann, Karl

2018-03-02

Chemical cell-surface engineering is a tool for modifying and altering cellular functions. Herein, we report the introduction of an antibiotic phenotype to the green alga Chlamydomonas reinhardtii by chemically modifying its cell surface. Flow cytometry and confocal microscopy studies demonstrated that a hybrid of the antibiotic vancomycin and a 4-hydroxyproline oligomer binds reversibly to the cell wall without affecting the viability or motility of the cells. The modified cells were used to inhibit bacterial growth of Gram-positive Bacillus subtilis cultures. Delivery of the antibiotic from the microalgae to the bacterial cells was verified by microscopy. Our studies provide compelling evidence that 1) chemical surface engineering constitutes a useful tool for the introduction of new, previously unknown functionality, and 2) living microalgae can serve as new platforms for drug delivery. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Deformation simulation of cells seeded on a collagen-GAG scaffold in a flow perfusion bioreactor using a sequential 3D CFD-elastostatics model.

PubMed

Jungreuthmayer, C; Jaasma, M J; Al-Munajjed, A A; Zanghellini, J; Kelly, D J; O'Brien, F J

2009-05-01

Tissue-engineered bone shows promise in meeting the huge demand for bone grafts caused by up to 4 million bone replacement procedures per year, worldwide. State-of-the-art bone tissue engineering strategies use flow perfusion bioreactors to apply biophysical stimuli to cells seeded on scaffolds and to grow tissue suitable for implantation into the patient's body. The aim of this study was to quantify the deformation of cells seeded on a collagen-GAG scaffold which was perfused by culture medium inside a flow perfusion bioreactor. Using a microCT scan of an unseeded collagen-GAG scaffold, a sequential 3D CFD-deformation model was developed. The wall shear stress and the hydrostatic wall pressure acting on the cells were computed through the use of a CFD simulation and fed into a linear elastostatics model in order to calculate the deformation of the cells. The model used numerically seeded cells of two common morphologies where cells are either attached flatly on the scaffold wall or bridging two struts of the scaffold. Our study showed that the displacement of the cells is primarily determined by the cell morphology. Although cells of both attachment profiles were subjected to the same mechanical load, cells bridging two struts experienced a deformation up to 500 times higher than cells only attached to one strut. As the scaffold's pore size determines both the mechanical load and the type of attachment, the design of an optimal scaffold must take into account the interplay of these two features and requires a design process that optimizes both parameters at the same time.

HOT CELL BUILDING, TRA632, INTERIOR. HOT CELL NO. 1 (THE ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

HOT CELL BUILDING, TRA-632, INTERIOR. HOT CELL NO. 1 (THE FIRST BUILT) IN LABORATORY 101. CAMERA FACES SOUTHEAST. SHIELDED OPERATING WINDOWS ARE ON LEFT (NORTH) SIDE. OBSERVATION WINDOW IS AT LEFT OF VIEW (ON WEST SIDE). PLASTIC COVERS SHROUD MASTER/SLAVE MANIPULATORS AT WINDOWS IN LEFT OF VIEW. NOTE MINERAL OIL RESERVOIR ABOVE "CELL 1" SIGN, INDICATING LEVEL OF THE FLUID INSIDE THE THICK WINDOWS. HOT CELL HAS BEVELED CORNER BECAUSE A SQUARED CORNER WOULD HAVE SUPPLIED UNNECESSARY SHIELDING. NOTE PUMICE BLOCK WALL AT LEFT OF VIEW. INL NEGATIVE NO. HD46-28-1. Mike Crane, Photographer, 2/2005 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

Systems and synthetic biology approaches to alter plant cell walls and reduce biomass recalcitrance.

PubMed

Kalluri, Udaya C; Yin, Hengfu; Yang, Xiaohan; Davison, Brian H

2014-12-01

Fine-tuning plant cell wall properties to render plant biomass more amenable to biofuel conversion is a colossal challenge. A deep knowledge of the biosynthesis and regulation of plant cell wall and a high-precision genome engineering toolset are the two essential pillars of efforts to alter plant cell walls and reduce biomass recalcitrance. The past decade has seen a meteoric rise in use of transcriptomics and high-resolution imaging methods resulting in fresh insights into composition, structure, formation and deconstruction of plant cell walls. Subsequent gene manipulation approaches, however, commonly include ubiquitous mis-expression of a single candidate gene in a host that carries an intact copy of the native gene. The challenges posed by pleiotropic and unintended changes resulting from such an approach are moving the field towards synthetic biology approaches. Synthetic biology builds on a systems biology knowledge base and leverages high-precision tools for high-throughput assembly of multigene constructs and pathways, precision genome editing and site-specific gene stacking, silencing and/or removal. Here, we summarize the recent breakthroughs in biosynthesis and remodelling of major secondary cell wall components, assess the impediments in obtaining a systems-level understanding and explore the potential opportunities in leveraging synthetic biology approaches to reduce biomass recalcitrance. Published 2014. This article is a U.S. Government work and is in the public domain in the USA. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Deconstructing a Plant Macromolecular Assembly: Chemical Architecture, Molecular Flexibility, And Mechanical Performance of Natural and Engineered Potato Suberins

PubMed Central

2015-01-01

Periderms present in plant barks are essential protective barriers to water diffusion, mechanical breakdown, and pathogenic invasion. They consist of densely packed layers of dead cells with cell walls that are embedded with suberin. Understanding the interplay of molecular structure, dynamics, and biomechanics in these cell wall-associated insoluble amorphous polymeric assemblies presents substantial investigative challenges. We report solid-state NMR coordinated with FT-IR and tensile strength measurements for periderms from native and wound-healing potatoes and from potatoes with genetically modified suberins. The analyses include the intact suberin aromatic–aliphatic polymer and cell-wall polysaccharides, previously reported soluble depolymerized transmethylation products, and undegraded residues including suberan. Wound-healing suberized potato cell walls, which are 2 orders of magnitude more permeable to water than native periderms, display a strikingly enhanced hydrophilic–hydrophobic balance, a degradation-resistant aromatic domain, and flexibility suggestive of an altered supramolecular organization in the periderm. Suppression of ferulate ester formation in suberin and associated wax remodels the periderm with more flexible aliphatic chains and abundant aromatic constituents that can resist transesterification, attenuates cooperative hydroxyfatty acid motions, and produces a mechanically compromised and highly water-permeable periderm. PMID:24502663

Deconstructing a plant macromolecular assembly: chemical architecture, molecular flexibility, and mechanical performance of natural and engineered potato suberins.

PubMed

Serra, Olga; Chatterjee, Subhasish; Figueras, Mercè; Molinas, Marisa; Stark, Ruth E

2014-03-10

Periderms present in plant barks are essential protective barriers to water diffusion, mechanical breakdown, and pathogenic invasion. They consist of densely packed layers of dead cells with cell walls that are embedded with suberin. Understanding the interplay of molecular structure, dynamics, and biomechanics in these cell wall-associated insoluble amorphous polymeric assemblies presents substantial investigative challenges. We report solid-state NMR coordinated with FT-IR and tensile strength measurements for periderms from native and wound-healing potatoes and from potatoes with genetically modified suberins. The analyses include the intact suberin aromatic-aliphatic polymer and cell-wall polysaccharides, previously reported soluble depolymerized transmethylation products, and undegraded residues including suberan. Wound-healing suberized potato cell walls, which are 2 orders of magnitude more permeable to water than native periderms, display a strikingly enhanced hydrophilic-hydrophobic balance, a degradation-resistant aromatic domain, and flexibility suggestive of an altered supramolecular organization in the periderm. Suppression of ferulate ester formation in suberin and associated wax remodels the periderm with more flexible aliphatic chains and abundant aromatic constituents that can resist transesterification, attenuates cooperative hydroxyfatty acid motions, and produces a mechanically compromised and highly water-permeable periderm.

Restricting lignin and enhancing sugar deposition in secondary cell walls enhances monomeric sugar release after low temperature ionic liquid pretreatment

DOE PAGES

Scullin, Chessa; Cruz, Alejandro G.; Chuang, Yi -De; ...

2015-07-04

Lignocellulosic biomass has the potential to be a major source of renewable sugar for biofuel production. Before enzymatic hydrolysis, biomass must first undergo a pretreatment step in order to be more susceptible to saccharification and generate high yields of fermentable sugars. Lignin, a complex, interlinked, phenolic polymer, associates with secondary cell wall polysaccharides, rendering them less accessible to enzymatic hydrolysis. Herein, we describe the analysis of engineered Arabidopsis lines where lignin biosynthesis was repressed in fiber tissues but retained in the vessels, and polysaccharide deposition was enhanced in fiber cells with little to no apparent negative impact on growth phenotype.

Restricting lignin and enhancing sugar deposition in secondary cell walls enhances monomeric sugar release after low temperature ionic liquid pretreatment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Scullin, Chessa; Cruz, Alejandro G.; Chuang, Yi -De

Lignocellulosic biomass has the potential to be a major source of renewable sugar for biofuel production. Before enzymatic hydrolysis, biomass must first undergo a pretreatment step in order to be more susceptible to saccharification and generate high yields of fermentable sugars. Lignin, a complex, interlinked, phenolic polymer, associates with secondary cell wall polysaccharides, rendering them less accessible to enzymatic hydrolysis. Herein, we describe the analysis of engineered Arabidopsis lines where lignin biosynthesis was repressed in fiber tissues but retained in the vessels, and polysaccharide deposition was enhanced in fiber cells with little to no apparent negative impact on growth phenotype.

Construction and histological analysis of a 3D human arterial wall model containing vasa vasorum using a layer-by-layer technique.

PubMed

Shima, Fumiaki; Narita, Hirokazu; Hiura, Ayami; Shimoda, Hiroshi; Akashi, Mitsuru

2017-03-01

There is considerable global demand for three-dimensional (3D) functional tissues which mimic our native organs and tissues for use as in vitro drug screening systems and in regenerative medicine. In particular, there has been an increasing number of patients who suffer from arterial diseases such as arteriosclerosis. As such, in vitro 3D arterial wall models that can evaluate the effects of novel medicines and a novel artificial graft for the treatment are required. In our previous study, we reported the rapid construction of 3D tissues by employing a layer-by-layer (LbL) technique and revealed their potential applications in the pharmaceutical fields and tissue engineering. In this study, we successfully constructed a 3D arterial wall model containing vasa vasorum by employing a LbL technique for the first time. The cells were coated with extracellular matrix nanofilms and seeded into a culture insert using a cell accumulation method. This model had a three-layered hierarchical structure: a fibroblast layer, a smooth muscle layer, and an endothelial layer, which resembled the native arterial wall. Our method could introduce vasa vasorum into a fibroblast layer in vitro and the 3D arterial wall model showed barrier function which was evaluated by immunostaining and transendothelial electrical resistance measurement. Furthermore, electron microscopy observations revealed that the vasa vasorum was composed of single-layered endothelial cells, and the endothelial tubes were surrounded by the basal lamina, which are known to promote maturation and stabilization in native blood capillaries. These models should be useful for tissue engineering, regenerative medicine, and pharmaceutical applications. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 814-823, 2017. © 2016 Wiley Periodicals, Inc.

Epigallocatechin gallate incorporation into lignin enhances the alkaline delignification and enzymatic saccharification of cell walls

PubMed Central

2012-01-01

Background Lignin is an integral component of the plant cell wall matrix but impedes the conversion of biomass into biofuels. The plasticity of lignin biosynthesis should permit the inclusion of new compatible phenolic monomers such as flavonoids into cell wall lignins that are consequently less recalcitrant to biomass processing. In the present study, epigallocatechin gallate (EGCG) was evaluated as a potential lignin bioengineering target for rendering biomass more amenable to processing for biofuel production. Results In vitro peroxidase-catalyzed polymerization experiments revealed that both gallate and pyrogallyl (B-ring) moieties in EGCG underwent radical cross-coupling with monolignols mainly by β–O–4-type cross-coupling, producing benzodioxane units following rearomatization reactions. Biomimetic lignification of maize cell walls with a 3:1 molar ratio of monolignols and EGCG permitted extensive alkaline delignification of cell walls (72 to 92%) that far exceeded that for lignified controls (44 to 62%). Alkali-insoluble residues from EGCG-lignified walls yielded up to 34% more glucose and total sugars following enzymatic saccharification than lignified controls. Conclusions It was found that EGCG readily copolymerized with monolignols to become integrally cross-coupled into cell wall lignins, where it greatly enhanced alkaline delignification and subsequent enzymatic saccharification. Improved delignification may be attributed to internal trapping of quinone-methide intermediates to prevent benzyl ether cross-linking of lignin to structural polysaccharides during lignification, and to the cleavage of ester intra-unit linkages within EGCG during pretreatment. Overall, our results suggest that apoplastic deposition of EGCG for incorporation into lignin would be a promising plant genetic engineering target for improving the delignification and saccharification of biomass crops. PMID:22889353

Scaffold-free, Human Mesenchymal Stem Cell-Based Tissue Engineered Blood Vessels.

PubMed

Jung, Youngmee; Ji, HaYeun; Chen, Zaozao; Fai Chan, Hon; Atchison, Leigh; Klitzman, Bruce; Truskey, George; Leong, Kam W

2015-10-12

Tissue-engineered blood vessels (TEBV) can serve as vascular grafts and may also play an important role in the development of organs-on-a-chip. Most TEBV construction involves scaffolding with biomaterials such as collagen gel or electrospun fibrous mesh. Hypothesizing that a scaffold-free TEBV may be advantageous, we constructed a tubular structure (1 mm i.d.) from aligned human mesenchymal cell sheets (hMSC) as the wall and human endothelial progenitor cell (hEPC) coating as the lumen. The burst pressure of the scaffold-free TEBV was above 200 mmHg after three weeks of sequential culture in a rotating wall bioreactor and perfusion at 6.8 dynes/cm(2). The interwoven organization of the cell layers and extensive extracellular matrix (ECM) formation of the hMSC-based TEBV resembled that of native blood vessels. The TEBV exhibited flow-mediated vasodilation, vasoconstriction after exposure to 1 μM phenylephrine and released nitric oxide in a manner similar to that of porcine femoral vein. HL-60 cells attached to the TEBV lumen after TNF-α activation to suggest a functional endothelium. This study demonstrates the potential of a hEPC endothelialized hMSC-based TEBV for drug screening.

Multi-scale mechanical response of freeze-dried collagen scaffolds for tissue engineering applications.

PubMed

Offeddu, Giovanni S; Ashworth, Jennifer C; Cameron, Ruth E; Oyen, Michelle L

2015-02-01

Tissue engineering has grown in the past two decades as a promising solution to unresolved clinical problems such as osteoarthritis. The mechanical response of tissue engineering scaffolds is one of the factors determining their use in applications such as cartilage and bone repair. The relationship between the structural and intrinsic mechanical properties of the scaffolds was the object of this study, with the ultimate aim of understanding the stiffness of the substrate that adhered cells experience, and its link to the bulk mechanical properties. Freeze-dried type I collagen porous scaffolds made with varying slurry concentrations and pore sizes were tested in a viscoelastic framework by macroindentation. Membranes made up of stacks of pore walls were indented using colloidal probe atomic force microscopy. It was found that the bulk scaffold mechanical response varied with collagen concentration in the slurry consistent with previous studies on these materials. Hydration of the scaffolds resulted in a more compliant response, yet lesser viscoelastic relaxation. Indentation of the membranes suggested that the material making up the pore walls remains unchanged between conditions, so that the stiffness of the scaffolds at the scale of seeded cells is unchanged; rather, it is suggested that thicker pore walls or more of these result in the increased moduli for the greater slurry concentration conditions. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

3D fiber deposited polymeric scaffolds for external auditory canal wall.

PubMed

Mota, Carlos; Milazzo, Mario; Panetta, Daniele; Trombi, Luisa; Gramigna, Vera; Salvadori, Piero A; Giannotti, Stefano; Bruschini, Luca; Stefanini, Cesare; Moroni, Lorenzo; Berrettini, Stefano; Danti, Serena

2018-05-07

The external auditory canal (EAC) is an osseocartilaginous structure extending from the auricle to the eardrum, which can be affected by congenital, inflammatory, and neoplastic diseases, thus reconstructive materials are needed. Current biomaterial-based approaches for the surgical reconstruction of EAC posterior wall still suffer from resorption (biological) and extrusion (synthetic). In this study, 3D fiber deposited scaffolds based on poly(ethylene oxide terephthalate)/poly(butylene terephthalate) were designed and fabricated to replace the EAC wall. Fiber diameter and scaffold porosity were optimized, leading to 200 ± 33 µm and 55% ± 5%, respectively. The mechanical properties were evaluated, resulting in a Young's modulus of 25.1 ± 7.0 MPa. Finally, the EAC scaffolds were tested in vitro with osteo-differentiated human mesenchymal stromal cells (hMSCs) with different seeding methods to produce homogeneously colonized replacements of interest for otologic surgery. This study demonstrated the fabrication feasibility of EAC wall scaffolds aimed to match several important requirements for biomaterial application to the ear under the Tissue Engineering paradigm, including shape, porosity, surface area, mechanical properties and favorable in vitro interaction with osteoinduced hMSCs. This study demonstrated the fabrication feasibility of outer ear canal wall scaffolds via additive manufacturing. Aimed to match several important requirements for biomaterial application to ear replacements under the Tissue Engineering paradigm, including shape, porosity and pore size, surface area, mechanical properties and favorable in vitro interaction with osteo-differentiated mesenchymal stromal cells.

Association Mapping of Cell Wall Synthesis Regulatory Genes and Cell Wall Quality in Switchgrass

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bartley, Laura; Wu, Y.; Zhu, L.

Inefficient conversion of biomass to biofuels is one of the main barriers for biofuel production from such materials. Approximately half of polysaccharides in biomass remain unused by typical biochemical conversion methods. Conversion efficiency is influenced by the composition and structure of cell walls of biomass. Grasses such as wheat, maize, and rice, as well as dedicated perennial bioenergy crops, like switchgrass, make up ~55% of biomass that can be produced in the United States. Grass cell walls have a different composition and patterning compared with dicotyledonous plants, including the well-studied model plant, Arabidopsis. This project identified genetic determinants of cellmore » wall composition in grasses using both naturally occurring genetic variation of switchgrass and gene network reconstruction and functional assays in rice. In addition, the project linked functional data in rice and other species to switchgrass improvement efforts through curation of the most abundant class of regulators in the switchgrass genome. Characterizing natural diversity of switchgrass for variation in cell wall composition and properties, also known as quality, provides an unbiased avenue for identifying biologically viable diversity in switchgrass cell walls. To characterizing natural diversity, this project generated cell wall composition and enzymatic deconstruction data for ~450 genotypes of the Switchgrass Southern Association Collection (SSAC), a diverse collection composed of 36 switchgrass accessions from the southern U.S. distribution of switchgrass. Comparing these data with other measures of cell wall quality for the same samples demonstrated the complementary nature of the diverse characterization platforms now being used for biomass characterization. Association of the composition data with ~3.2K single nucleotide variant markers identified six significant single nucleotide variant markers co-associated with digestibility and another compositional trait. These markers might be used to select switchgrass genotypes with improved composition in breeding programs for biofuel and forage production. Because the SSAC continues to be characterized by collaborators in the bioenergy community, the data generated will be used to identify additional markers in higher resolution genotyping data to approach identifying the genes and alleles that cause natural variation in switchgrass cell wall quality. For example, these markers can be surveyed in the 2100-member Oklahoma Southern and Northern Lowland switchgrass collections that this project also characterized. An orthogonal approach to biodiversity studies, using comparative functional genomics permits systematic querying of how much regulatory information is likely to be transferable from dicots to grasses and use of accumulated functional genomics resources for better-characterized grass species, such as rice, itself a biomass source in global agriculture and in certain regions. The project generated and tested a number of specific hypotheses regarding cell wall transcription factors and enzymes of grasses. To aid identification of cell wall regulators, the project assembled a novel, highdepth and -quality gene association network using a general linearized model scoring system to combine rice gene network data. Using known or putative orthologs of Arabidopsis cell wall biosynthesis genes and regulators, the project pulled from this network a cell wall sub-network that includes 96 transcription factors. Reverse genetics of a co-ortholog of the Arabidopsis MYB61 transcription factor in rice revealed that this regulatory node has evolved the ability to regulate grass-specific cell wall synthesis enzymes. A transcription factor with such activity has not been previously characterized to our knowledge, representing a major conclusion of this work. Changes in gene expression in a protoplast-based assay demonstrated positive or negative roles in cell wall regulation for eleven other transcription factors from the rice gene network. Eight of fifteen (53%) of these have not previously been examined for this function. Some of these may represent novel grass-diverged cell wall regulators, while others are likely to have this function across angiosperms. A parallel effort of this project to expand knowledge of enzymes that have evolved to function in grass cell wall synthesis, revealed that a grass-diverged enzyme in rice, OsAT 5, ferulates monolignols that are naturally incorporated into grass cell walls. This finding opens potential natural selection avenues for improving biomass composition for downstream processing by weak base pretreatment. Thus, this project has significantly expanded knowledge of cell wall synthesis and regulation in rice, information that can be used in reverse genetics and synthetic biology approaches to re-engineer cell walls for improved production of biofuel and high-value products. To lay the foundation for translating these results directly for switchgrass improvement, the project employed a comparative phylogenetic analysis of the major group of cell wall transcription factors that have been found to function in cell wall regulation, the R 2R 3 MYBs. This analysis concluded that known cell wall regulators are largely conserved across switchgrass, rice, maize, poplar, and Arabidopsis. This interpretation is also largely consistent with the gene network analysis described above, though both approaches provide evidence that some co-orthologs of Arabidopsis regulators have diminished or increased in importance based on gene expression patterns. Also, several clades containing dicot cell wall regulators have expanded, consistent with the evolution of new cell wall regulators. This latter result is supported by functional analysis of the R 2R 3 MYB protein SWAM 1 in a collaboration between this project and the DOE-funded group of Dr. S. Hazen at the University of Massachusettes. The curation of the switchgrass genome through this project provides specific targets for future engineering of switchgrass cell wall regulation and may also facilitate identification of regulators that underlie the molecular markers that are genetically linked to differences in cell wall quality. With the goal of spurring further research and technological developments in lignocellulosic biofuel production, this work has been communicated to the bioenergy and cell wall communities though various presentations and publications. To date, three manuscripts have been published, two others are near to publication, three others are in an advanced state, and two to four more are likely to be written based on analyses still in progress. In addition, project participants have presented thirteen posters and talks at regional, national, and international meetings about aspects of this project. In sum, the work supported by this funding has made and communicated significant progress in identifying the genes that grasses use for cell wall synthesis and regulation, information that will be used by project participants and others to improve the efficiency of conversion of lignocellulosic biomass to biofuels.« less

Transgenic Plant-Produced Hydrolytic Enzymes and the Potential of Insect Gut-Derived Hydrolases for Biofuels

PubMed Central

Willis, Jonathan D.; Mazarei, Mitra; Stewart, C. Neal

2016-01-01

Various perennial C4 grass species have tremendous potential for use as lignocellulosic biofuel feedstocks. Currently available grasses require costly pre-treatment and exogenous hydrolytic enzyme application to break down complex cell wall polymers into sugars that can then be fermented into ethanol. It has long been hypothesized that engineered feedstock production of cell wall degrading (CWD) enzymes would be an efficient production platform for of exogenous hydrolytic enzymes. Most research has focused on plant overexpression of CWD enzyme-coding genes from free-living bacteria and fungi that naturally break down plant cell walls. Recently, it has been found that insect digestive tracts harbor novel sources of lignocellulolytic biocatalysts that might be exploited for biofuel production. These CWD enzyme genes can be located in the insect genomes or in symbiotic microbes. When CWD genes are transformed into plants, negative pleiotropic effects are possible such as unintended cell wall digestion. The use of codon optimization along with organelle and tissue specific targeting improves CWD enzyme yields. The literature teaches several important lessons on strategic deployment of CWD genes in transgenic plants, which is the focus of this review. PMID:27303411

Strategies for the production of cell wall-deconstructing enzymes in lignocellulosic biomass and their utilization for biofuel production

DOE PAGES

Park, Sang -Hyuck; Ong, Rebecca Garlock; Sticklen, Mariam

2015-12-02

Microbial cell wall-deconstructing enzymes are widely used in the food, wine, pulp and paper, textile, and detergent industries and will be heavily utilized by cellulosic biorefineries in the production of fuels and chemicals. Due to their ability to use freely available solar energy, genetically engineered bioenergy crops provide an attractive alternative to microbial bioreactors for the production of cell wall-deconstructing enzymes. This review article summarizes the efforts made within the last decade on the production of cell wall-deconstructing enzymes in planta for use in the deconstruction of lignocellulosic biomass. A number of strategies have been employed to increase enzyme yieldsmore » and limit negative impacts on plant growth and development including targeting heterologous enzymes into specific subcellular compartments using signal peptides, using tissue-specific or inducible promoters to limit the expression of enzymes to certain portions of the plant or certain times, and fusion of amplification sequences upstream of the coding region to enhance expression. As a result, we also summarize methods that have been used to access and maintain activity of plant-generated enzymes when used in conjunction with thermochemical pretreatments for the production of lignocellulosic biofuels.« less

A Novel Bioreactor System for the Assessment of Endothelialization on Deformable Surfaces

PubMed Central

Bachmann, Björn J.; Bernardi, Laura; Loosli, Christian; Marschewski, Julian; Perrini, Michela; Ehrbar, Martin; Ermanni, Paolo; Poulikakos, Dimos; Ferrari, Aldo; Mazza, Edoardo

2016-01-01

The generation of a living protective layer at the luminal surface of cardiovascular devices, composed of an autologous functional endothelium, represents the ideal solution to life-threatening, implant-related complications in cardiovascular patients. The initial evaluation of engineering strategies fostering endothelial cell adhesion and proliferation as well as the long-term tissue homeostasis requires in vitro testing in environmental model systems able to recapitulate the hemodynamic conditions experienced at the blood-to-device interface of implants as well as the substrate deformation. Here, we introduce the design and validation of a novel bioreactor system which enables the long-term conditioning of human endothelial cells interacting with artificial materials under dynamic combinations of flow-generated wall shear stress and wall deformation. The wall shear stress and wall deformation values obtained encompass both the physiological and supraphysiological range. They are determined through separate actuation systems which are controlled based on validated computational models. In addition, we demonstrate the good optical conductivity of the system permitting online monitoring of cell activities through live-cell imaging as well as standard biochemical post-processing. Altogether, the bioreactor system defines an unprecedented testing hub for potential strategies toward the endothelialization or re-endothelialization of target substrates. PMID:27941901

Strategies for the production of cell wall-deconstructing enzymes in lignocellulosic biomass and their utilization for biofuel production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Sang -Hyuck; Ong, Rebecca Garlock; Sticklen, Mariam

Microbial cell wall-deconstructing enzymes are widely used in the food, wine, pulp and paper, textile, and detergent industries and will be heavily utilized by cellulosic biorefineries in the production of fuels and chemicals. Due to their ability to use freely available solar energy, genetically engineered bioenergy crops provide an attractive alternative to microbial bioreactors for the production of cell wall-deconstructing enzymes. This review article summarizes the efforts made within the last decade on the production of cell wall-deconstructing enzymes in planta for use in the deconstruction of lignocellulosic biomass. A number of strategies have been employed to increase enzyme yieldsmore » and limit negative impacts on plant growth and development including targeting heterologous enzymes into specific subcellular compartments using signal peptides, using tissue-specific or inducible promoters to limit the expression of enzymes to certain portions of the plant or certain times, and fusion of amplification sequences upstream of the coding region to enhance expression. As a result, we also summarize methods that have been used to access and maintain activity of plant-generated enzymes when used in conjunction with thermochemical pretreatments for the production of lignocellulosic biofuels.« less

Biodegradable scaffold with built-in vasculature for organ-on-a-chip engineering and direct surgical anastomosis.

PubMed

Zhang, Boyang; Montgomery, Miles; Chamberlain, M Dean; Ogawa, Shinichiro; Korolj, Anastasia; Pahnke, Aric; Wells, Laura A; Massé, Stéphane; Kim, Jihye; Reis, Lewis; Momen, Abdul; Nunes, Sara S; Wheeler, Aaron R; Nanthakumar, Kumaraswamy; Keller, Gordon; Sefton, Michael V; Radisic, Milica

2016-06-01

We report the fabrication of a scaffold (hereafter referred to as AngioChip) that supports the assembly of parenchymal cells on a mechanically tunable matrix surrounding a perfusable, branched, three-dimensional microchannel network coated with endothelial cells. The design of AngioChip decouples the material choices for the engineered vessel network and for cell seeding in the parenchyma, enabling extensive remodelling while maintaining an open-vessel lumen. The incorporation of nanopores and micro-holes in the vessel walls enhances permeability, and permits intercellular crosstalk and extravasation of monocytes and endothelial cells on biomolecular stimulation. We also show that vascularized hepatic tissues and cardiac tissues engineered by using AngioChips process clinically relevant drugs delivered through the vasculature, and that millimetre-thick cardiac tissues can be engineered in a scalable manner. Moreover, we demonstrate that AngioChip cardiac tissues implanted with direct surgical anastomosis to the femoral vessels of rat hindlimbs establish immediate blood perfusion.

Biodegradable scaffold with built-in vasculature for organ-on-a-chip engineering and direct surgical anastomosis

NASA Astrophysics Data System (ADS)

Zhang, Boyang; Montgomery, Miles; Chamberlain, M. Dean; Ogawa, Shinichiro; Korolj, Anastasia; Pahnke, Aric; Wells, Laura A.; Massé, Stéphane; Kim, Jihye; Reis, Lewis; Momen, Abdul; Nunes, Sara S.; Wheeler, Aaron R.; Nanthakumar, Kumaraswamy; Keller, Gordon; Sefton, Michael V.; Radisic, Milica

2016-06-01

We report the fabrication of a scaffold (hereafter referred to as AngioChip) that supports the assembly of parenchymal cells on a mechanically tunable matrix surrounding a perfusable, branched, three-dimensional microchannel network coated with endothelial cells. The design of AngioChip decouples the material choices for the engineered vessel network and for cell seeding in the parenchyma, enabling extensive remodelling while maintaining an open-vessel lumen. The incorporation of nanopores and micro-holes in the vessel walls enhances permeability, and permits intercellular crosstalk and extravasation of monocytes and endothelial cells on biomolecular stimulation. We also show that vascularized hepatic tissues and cardiac tissues engineered by using AngioChips process clinically relevant drugs delivered through the vasculature, and that millimetre-thick cardiac tissues can be engineered in a scalable manner. Moreover, we demonstrate that AngioChip cardiac tissues implanted with direct surgical anastomosis to the femoral vessels of rat hindlimbs establish immediate blood perfusion.

Accessibility and contribution to glucan masking of natural and genetically tagged versions of yeast wall protein 1 of Candida albicans

PubMed Central

2018-01-01

Yeast wall protein 1 (Ywp1) is an abundant glycoprotein of the cell wall of the yeast form of Candida albicans, the most prevalent fungal pathogen of humans. Antibodies that bind to the polypeptide backbone of isolated Ywp1 show little binding to intact yeast cells, presumably because the Ywp1 epitopes are masked by the polysaccharides of the mannoproteins that form the outer layer of the cell wall. Rare cells do exhibit much greater anti-Ywp1 binding, however, and one of these was isolated and characterized. No differences were seen in its Ywp1, but it exhibited greater adhesiveness, sensitivity to wall perturbing agents, and exposure of its underlying β-1,3-glucan layer to external antibodies. The molecular basis for this greater epitope accessibility has not been determined, but has facilitated exploration of how these properties change as a function of cell growth and morphology. In addition, previously engineered strains with reduced quantities of Ywp1 in their cell walls were also found to have greater β-1,3-glucan exposure, indicating that Ywp1 itself contributes to the masking of wall epitopes, which may be important for understanding the anti-adhesive effect of Ywp1. Ectopic production of Ywp1 by hyphae, which reduces the adhesivity of these filamentous forms of C. albicans, was similarly found to reduce exposure of the β-1,3-glucan in their walls. To monitor Ywp1 in the cell wall irrespective of its accessibility, green fluorescent protein (Gfp) was genetically inserted into wall-anchored Ywp1 using a bifunctional cassette that also allowed production from a single transfection of a soluble, anchor-free version. The wall-anchored Ywp1-Gfp-Ywp1 accumulated in the wall of the yeast forms but not hyphae, and appeared to have properties similar to native Ywp1, including its adhesion-inhibiting effect. Some pseudohyphal walls also detectably accumulated this probe. Strains of C. albicans with tandem hemagglutinin (HA) epitopes inserted into wall-anchored Ywp1 were previously created by others, and were further explored here. As above, rare cells with much greater accessibility of the HA epitopes were isolated, and also found to exhibit greater exposure of Ywp1 and β-1,3-glucan. The placement of the HA cassette inhibited the normal N-glycosylation and propeptide cleavage of Ywp1, but the wall-anchored Ywp1-HA-Ywp1 still accumulated in the cell wall of yeast forms. Bifunctional transformation cassettes were used to additionally tag these molecules with Gfp, generating soluble Ywp1-HA-Gfp and wall-anchored Ywp1-HA-Gfp-Ywp1 molecules. The former revealed unexpected electrophoretic properties caused by the HA insertion, while the latter further highlighted differences between the presence of a tagged Ywp1 molecule (as revealed by Gfp fluorescence) and its accessibility in the cell wall to externally applied antibodies specific for HA, Gfp and Ywp1, with accessibility being greatest in the rapidly expanding walls of budding daughter cells. These strains and results increase our understanding of cell wall properties and how C. albicans masks itself from recognition by the human immune system. PMID:29329339

Accessibility and contribution to glucan masking of natural and genetically tagged versions of yeast wall protein 1 of Candida albicans.

PubMed

Granger, Bruce L

2018-01-01

Yeast wall protein 1 (Ywp1) is an abundant glycoprotein of the cell wall of the yeast form of Candida albicans, the most prevalent fungal pathogen of humans. Antibodies that bind to the polypeptide backbone of isolated Ywp1 show little binding to intact yeast cells, presumably because the Ywp1 epitopes are masked by the polysaccharides of the mannoproteins that form the outer layer of the cell wall. Rare cells do exhibit much greater anti-Ywp1 binding, however, and one of these was isolated and characterized. No differences were seen in its Ywp1, but it exhibited greater adhesiveness, sensitivity to wall perturbing agents, and exposure of its underlying β-1,3-glucan layer to external antibodies. The molecular basis for this greater epitope accessibility has not been determined, but has facilitated exploration of how these properties change as a function of cell growth and morphology. In addition, previously engineered strains with reduced quantities of Ywp1 in their cell walls were also found to have greater β-1,3-glucan exposure, indicating that Ywp1 itself contributes to the masking of wall epitopes, which may be important for understanding the anti-adhesive effect of Ywp1. Ectopic production of Ywp1 by hyphae, which reduces the adhesivity of these filamentous forms of C. albicans, was similarly found to reduce exposure of the β-1,3-glucan in their walls. To monitor Ywp1 in the cell wall irrespective of its accessibility, green fluorescent protein (Gfp) was genetically inserted into wall-anchored Ywp1 using a bifunctional cassette that also allowed production from a single transfection of a soluble, anchor-free version. The wall-anchored Ywp1-Gfp-Ywp1 accumulated in the wall of the yeast forms but not hyphae, and appeared to have properties similar to native Ywp1, including its adhesion-inhibiting effect. Some pseudohyphal walls also detectably accumulated this probe. Strains of C. albicans with tandem hemagglutinin (HA) epitopes inserted into wall-anchored Ywp1 were previously created by others, and were further explored here. As above, rare cells with much greater accessibility of the HA epitopes were isolated, and also found to exhibit greater exposure of Ywp1 and β-1,3-glucan. The placement of the HA cassette inhibited the normal N-glycosylation and propeptide cleavage of Ywp1, but the wall-anchored Ywp1-HA-Ywp1 still accumulated in the cell wall of yeast forms. Bifunctional transformation cassettes were used to additionally tag these molecules with Gfp, generating soluble Ywp1-HA-Gfp and wall-anchored Ywp1-HA-Gfp-Ywp1 molecules. The former revealed unexpected electrophoretic properties caused by the HA insertion, while the latter further highlighted differences between the presence of a tagged Ywp1 molecule (as revealed by Gfp fluorescence) and its accessibility in the cell wall to externally applied antibodies specific for HA, Gfp and Ywp1, with accessibility being greatest in the rapidly expanding walls of budding daughter cells. These strains and results increase our understanding of cell wall properties and how C. albicans masks itself from recognition by the human immune system.

Modeling of flow-induced shear stress applied on 3D cellular scaffolds: Implications for vascular tissue engineering.

PubMed

Lesman, Ayelet; Blinder, Yaron; Levenberg, Shulamit

2010-02-15

Novel tissue-culture bioreactors employ flow-induced shear stress as a means of mechanical stimulation of cells. We developed a computational fluid dynamics model of the complex three-dimensional (3D) microstructure of a porous scaffold incubated in a direct perfusion bioreactor. Our model was designed to predict high shear-stress values within the physiological range of those naturally sensed by vascular cells (1-10 dyne/cm(2)), and will thereby provide suitable conditions for vascular tissue-engineering experiments. The model also accounts for cellular growth, which was designed as an added cell layer grown on all scaffold walls. Five model variants were designed, with geometric differences corresponding to cell-layer thicknesses of 0, 50, 75, 100, and 125 microm. Four inlet velocities (0.5, 1, 1.5, and 2 cm/s) were applied to each model. Wall shear-stress distribution and overall pressure drop calculations were then used to characterize the relation between flow rate, shear stress, cell-layer thickness, and pressure drop. The simulations showed that cellular growth within 3D scaffolds exposes cells to elevated shear stress, with considerably increasing average values in correlation to cell growth and inflow velocity. Our results provide in-depth analysis of the microdynamic environment of cells cultured within 3D environments, and thus provide advanced control over tissue development in vitro. 2009 Wiley Periodicals, Inc.

A&M. TAN607. Special service cubicle (hot cell). Details include Zpipe ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

A&M. TAN-607. Special service cubicle (hot cell). Details include Z-pipe and stepped plug penetrations through shielding wall. Ralph M. Parsons 902-3-ANP-607-A116. Date: December 1952. Approved by INEEL Classification Office for public release. INEEL index code no. 034-0607-693-106767 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

Tissue Engineering Under Microgravity Conditions-Use of Stem Cells and Specialized Cells.

PubMed

Grimm, Daniela; Egli, Marcel; Krüger, Marcus; Riwaldt, Stefan; Corydon, Thomas J; Kopp, Sascha; Wehland, Markus; Wise, Petra; Infanger, Manfred; Mann, Vivek; Sundaresan, Alamelu

2018-03-29

Experimental cell research studying three-dimensional (3D) tissues in space and on Earth using new techniques to simulate microgravity is currently a hot topic in Gravitational Biology and Biomedicine. This review will focus on the current knowledge of the use of stem cells and specialized cells for tissue engineering under simulated microgravity conditions. We will report on recent advancements in the ability to construct 3D aggregates from various cell types using devices originally created to prepare for spaceflights such as the random positioning machine (RPM), the clinostat, or the NASA-developed rotating wall vessel (RWV) bioreactor, to engineer various tissues such as preliminary vessels, eye tissue, bone, cartilage, multicellular cancer spheroids, and others from different cells. In addition, stem cells had been investigated under microgravity for the purpose to engineer adipose tissue, cartilage, or bone. Recent publications have discussed different changes of stem cells when exposed to microgravity and the relevant pathways involved in these biological processes. Tissue engineering in microgravity is a new technique to produce organoids, spheroids, or tissues with and without scaffolds. These 3D aggregates can be used for drug testing studies or for coculture models. Multicellular tumor spheroids may be interesting for radiation experiments in the future and to reduce the need for in vivo experiments. Current achievements using cells from patients engineered on the RWV or on the RPM represent an important step in the advancement of techniques that may be applied in translational Regenerative Medicine.

Plant cell wall glycosyltransferases: High-throughput recombinant expression screening and general requirements for these challenging enzymes

DOE PAGES

Welner, Ditte Hededam; Shin, David; Tomaleri, Giovani P.; ...

2017-06-09

Molecular characterization of plant cell wall glycosyltransferases is a critical step towards understanding the biosynthesis of the complex plant cell wall, and ultimately for efficient engineering of biofuel and agricultural crops. The majority of these enzymes have proven very difficult to obtain in the needed amount and purity for such molecular studies, and recombinant cell wall glycosyltransferase production efforts have largely failed. A daunting number of strategies can be employed to overcome this challenge, including optimization of DNA and protein sequences, choice of expression organism, expression conditions, coexpression partners, purification methods, and optimization of protein solubility and stability. Hence researchersmore » are presented with thousands of potential conditions to test. Ultimately, the subset of conditions that will be sampled depends on practical considerations and prior knowledge of the enzyme(s) being studied. We have developed a rational approach to this process. We devise a pipeline comprising in silico selection of targets and construct design, and high-throughput expression screening, target enrichment, and hit identification. We have applied this pipeline to a test set of Arabidopsis thaliana cell wall glycosyltransferases known to be challenging to obtain in soluble form, as well as to a library of cell wall glycosyltransferases from other plants including agricultural and biofuel crops. The screening results suggest that recombinant cell wall glycosyltransferases in general have a very low soluble: Insoluble ratio in lysates from heterologous expression cultures, and that co-expression of chaperones as well as lysis buffer optimization can increase this ratio. We have applied the identified preferred conditions to Reversibly Glycosylated Polypeptide 1 from Arabidopsis thaliana, and processed this enzyme to near-purity in unprecedented milligram amounts. The obtained preparation of Reversibly Glycosylated Polypeptide 1 has the expected arabinopyranose mutase and autoglycosylation activities.This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.« less

Lignin biosynthesis perturbations affect secondary cell wall composition and saccharification yield in Arabidopsis thaliana

PubMed Central

2013-01-01

Background Second-generation biofuels are generally produced from the polysaccharides in the lignocellulosic plant biomass, mainly cellulose. However, because cellulose is embedded in a matrix of other polysaccharides and lignin, its hydrolysis into the fermentable glucose is hampered. The senesced inflorescence stems of a set of 20 Arabidopsis thaliana mutants in 10 different genes of the lignin biosynthetic pathway were analyzed for cell wall composition and saccharification yield. Saccharification models were built to elucidate which cell wall parameters played a role in cell wall recalcitrance. Results Although lignin is a key polymer providing the strength necessary for the plant’s ability to grow upward, a reduction in lignin content down to 64% of the wild-type level in Arabidopsis was tolerated without any obvious growth penalty. In contrast to common perception, we found that a reduction in lignin was not compensated for by an increase in cellulose, but rather by an increase in matrix polysaccharides. In most lignin mutants, the saccharification yield was improved by up to 88% cellulose conversion for the cinnamoyl-coenzyme A reductase1 mutants under pretreatment conditions, whereas the wild-type cellulose conversion only reached 18%. The saccharification models and Pearson correlation matrix revealed that the lignin content was the main factor determining the saccharification yield. However, also lignin composition, matrix polysaccharide content and composition, and, especially, the xylose, galactose, and arabinose contents influenced the saccharification yield. Strikingly, cellulose content did not significantly affect saccharification yield. Conclusions Although the lignin content had the main effect on saccharification, also other cell wall factors could be engineered to potentially increase the cell wall processability, such as the galactose content. Our results contribute to a better understanding of the effect of lignin perturbations on plant cell wall composition and its influence on saccharification yield, and provide new potential targets for genetic improvement. PMID:23622268

Plant cell wall glycosyltransferases: High-throughput recombinant expression screening and general requirements for these challenging enzymes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Welner, Ditte Hededam; Shin, David; Tomaleri, Giovani P.

Molecular characterization of plant cell wall glycosyltransferases is a critical step towards understanding the biosynthesis of the complex plant cell wall, and ultimately for efficient engineering of biofuel and agricultural crops. The majority of these enzymes have proven very difficult to obtain in the needed amount and purity for such molecular studies, and recombinant cell wall glycosyltransferase production efforts have largely failed. A daunting number of strategies can be employed to overcome this challenge, including optimization of DNA and protein sequences, choice of expression organism, expression conditions, coexpression partners, purification methods, and optimization of protein solubility and stability. Hence researchersmore » are presented with thousands of potential conditions to test. Ultimately, the subset of conditions that will be sampled depends on practical considerations and prior knowledge of the enzyme(s) being studied. We have developed a rational approach to this process. We devise a pipeline comprising in silico selection of targets and construct design, and high-throughput expression screening, target enrichment, and hit identification. We have applied this pipeline to a test set of Arabidopsis thaliana cell wall glycosyltransferases known to be challenging to obtain in soluble form, as well as to a library of cell wall glycosyltransferases from other plants including agricultural and biofuel crops. The screening results suggest that recombinant cell wall glycosyltransferases in general have a very low soluble: Insoluble ratio in lysates from heterologous expression cultures, and that co-expression of chaperones as well as lysis buffer optimization can increase this ratio. We have applied the identified preferred conditions to Reversibly Glycosylated Polypeptide 1 from Arabidopsis thaliana, and processed this enzyme to near-purity in unprecedented milligram amounts. The obtained preparation of Reversibly Glycosylated Polypeptide 1 has the expected arabinopyranose mutase and autoglycosylation activities.This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.« less

[Histocompatibility of nano-hydroxyapatite/poly-co-glycolic acid tissue engineering bone modified by mesenchymal stem cells with vascular endothelial frowth factor].

PubMed

Zhang, Minglei; Wang, Dapeng; Yin, Ruofeng

2015-10-06

To explorec Histocompatibility of nano-hydroxyapatite/poly-co-glycolic acid tissue engineering bone modified by mesenchymal stem cells with vascular endothelial frowth factor transinfected. Rat bone marrow mesenchymal stem cells (BMSCs) was separated, using BMSCs as target cells, and then vascular endothelial growth factor (VEGF) gene was transfected. Composite bone marrow mesenchymal stem cells and cells transfected with nano-hydroxyapatite (HA)/polylactic-co-glycolic acid (PLGA). The composition of cell and scaffold was observed. The blank plasmid transfection was 39.1%, 40.1% in VEGF group. The cell adhesion and growth was found on the scaffold pore wall after 5 days, and the number of adherent cells in the nano-HA/PLGA composite scaffold material basically had no significant difference in both. Although the nano-HA/PLGA scaffold material is still not fully meet the requirements of the matrix material for bone tissue engineering, but good biocompatibility, structure is its rich microporous satisfaction in material mechanics, toughening, enhanced obviously. Composition scaffold with BMSCs transfected by VEGF plasmid, the ability of angiogenesis is promoted.

Single walled carbon nanotube composites for bone tissue engineering.

PubMed

Gupta, Ashim; Woods, Mia D; Illingworth, Kenneth David; Niemeier, Ryan; Schafer, Isaac; Cady, Craig; Filip, Peter; El-Amin, Saadiq F

2013-09-01

The purpose of this study was to develop single walled carbon nanotubes (SWCNT) and poly lactic-co-glycolic acid (PLAGA) composites for orthopedic applications and to evaluate the interaction of human stem cells (hBMSCs) and osteoblasts (MC3T3-E1 cells) via cell growth, proliferation, gene expression, extracellular matrix production and mineralization. PLAGA and SWCNT/PLAGA composites were fabricated with various amounts of SWCNT (5, 10, 20, 40, and 100 mg), characterized and degradation studies were performed. Cells were seeded and cell adhesion/morphology, growth/survival, proliferation and gene expression analysis were performed to evaluate biocompatibility. Imaging studies demonstrated uniform incorporation of SWCNT into the PLAGA matrix and addition of SWCNT did not affect the degradation rate. Imaging studies revealed that MC3T3-E1 and hBMSCs cells exhibited normal, non-stressed morphology on the composites and all were biocompatible. Composites with 10 mg SWCNT resulted in highest rate of cell proliferation (p < 0.05) among all composites. Gene expression of alkaline phosphatase, collagen I, osteocalcin, osteopontin, Runx-2, and Bone Sialoprotein was observed on all composites. In conclusion, SWCNT/PLAGA composites imparted beneficial cellular growth capabilities and gene expression, and mineralization abilities were well established. These results demonstrate the potential of SWCNT/PLAGA composites for musculoskeletal regeneration and bone tissue engineering (BTE) and are promising for orthopedic applications. Copyright © 2013 Orthopaedic Research Society.

SPERTI Instrument Cell Building (PER606) elevation; plan of Guard House ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

SPERT-I Instrument Cell Building (PER-606) elevation; plan of Guard House (PER-607); elevations for Pit Building (PER-605) southwest, southeast, and northeast sides. Earthen shield is mounded between back wall of Instrument Cell Building and the southwest elevation of Pit Building. Detail of filtered louver in door of Instrument Cell Building. Idaho Operations Office PER-605-IDO-3. INEEL index no. 761-0605-00-396-109183 - Idaho National Engineering Laboratory, SPERT-I & Power Burst Facility Area, Scoville, Butte County, ID

Micro-engineered first wall tungsten armor for high average power laser fusion energy systems

NASA Astrophysics Data System (ADS)

Sharafat, Shahram; Ghoniem, Nasr M.; Anderson, Michael; Williams, Brian; Blanchard, Jake; Snead, Lance; HAPL Team

2005-12-01

The high average power laser program is developing an inertial fusion energy demonstration power reactor with a solid first wall chamber. The first wall (FW) will be subject to high energy density radiation and high doses of high energy helium implantation. Tungsten has been identified as the candidate material for a FW armor. The fundamental concern is long term thermo-mechanical survivability of the armor against the effects of high temperature pulsed operation and exfoliation due to the retention of implanted helium. Even if a solid tungsten armor coating would survive the high temperature cyclic operation with minimal failure, the high helium implantation and retention would result in unacceptable material loss rates. Micro-engineered materials, such as castellated structures, plasma sprayed nano-porous coatings and refractory foams are suggested as a first wall armor material to address these fundamental concerns. A micro-engineered FW armor would have to be designed with specific geometric features that tolerate high cyclic heating loads and recycle most of the implanted helium without any significant failure. Micro-engineered materials are briefly reviewed. In particular, plasma-sprayed nano-porous tungsten and tungsten foams are assessed for their potential to accommodate inertial fusion specific loads. Tests show that nano-porous plasma spray coatings can be manufactured with high permeability to helium gas, while retaining relatively high thermal conductivities. Tungsten foams where shown to be able to overcome thermo-mechanical loads by cell rotation and deformation. Helium implantation tests have shown, that pulsed implantation and heating releases significant levels of implanted helium. Helium implantation and release from tungsten was modeled using an expanded kinetic rate theory, to include the effects of pulsed implantations and thermal cycles. Although, significant challenges remain micro-engineered materials are shown to constitute potential candidate FW armor materials.

Enhancing digestibility and ethanol yield of Populus wood via expression of an engineered monolignol 4-O-methyltransferase

PubMed Central

Cai, Yuanheng; Zhang, Kewei; Kim, Hoon; Hou, Guichuan; Zhang, Xuebin; Yang, Huijun; Feng, Huan; Miller, Lisa; Ralph, John; Liu, Chang-Jun

2016-01-01

Producing cellulosic biofuels and bio-based chemicals from woody biomass is impeded by the presence of lignin polymer in the plant cell wall. Manipulating the monolignol biosynthetic pathway offers a promising approach to improved processability, but often impairs plant growth and development. Here, we show that expressing an engineered 4-O-methyltransferase that chemically modifies the phenolic moiety of lignin monomeric precursors, thus preventing their incorporation into the lignin polymer, substantially alters hybrid aspens' lignin content and structure. Woody biomass derived from the transgenic aspens shows a 62% increase in the release of simple sugars and up to a 49% increase in the yield of ethanol when the woody biomass is subjected to enzymatic digestion and yeast-mediated fermentation. Moreover, the cell wall structural changes do not affect growth and biomass production of the trees. Our study provides a useful strategy for tailoring woody biomass for bio-based applications. PMID:27349324

Enhancing digestibility and ethanol yield of Populus wood via expression of an engineered monolignol 4-O-methyltransferase

DOE PAGES

Cai, Yuanheng; Zhang, Kewei; Kim, Hoon; ...

2016-06-28

Producing cellulosic biofuels and bio-based chemicals from woody biomass is impeded by the presence of lignin polymer in the plant cell wall. Manipulating the monolignol biosynthetic pathway offers a promising approach to improved processability, but often impairs plant growth and development. Here, we show that expressing an engineered 4-O-methyltransferase that chemically modifies the phenolic moiety of lignin monomeric precursors, thus preventing their incorporation into the lignin polymer, substantially alters hybrid aspens’ lignin content and structure. Woody biomass derived from the transgenic aspens shows a 62% increase in the release of simple sugars and up to a 49% increase in themore » yield of ethanol when the woody biomass is subjected to enzymatic digestion and yeast-mediated fermentation. Furthermore, the cell wall structural changes do not affect growth and biomass production of the trees. Our study provides a useful strategy for tailoring woody biomass for bio-based applications.« less

Use and engineering of efflux pumps for the export of olefins in microbes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mukhopadhyay, Aindrila

2016-07-14

The scope of the project is to investigate efflux pump systems in engineered host microorganisms, such as E. coli, and develop a pump engineered to export a target compound. To initiate the project in coordination with other TOTAL driven projects, the first target compound to be studied was 1-hexene. However, we were investigating other chemicals as Styrene. The main goal of the project was to generate a set of optimized efflux pump systems for microorganisms (E. coli and Streptomyces or other host) engineered to contain biosynthetic pathways to export large titers of target compounds that are toxic (or accumulate andmore » push back biosynthesis) to the host cell. An optimized microbial host will utilize specific and efficient cell wall located pumps to extrude harmful target compounds and enable greater production of these compounds.« less

Engineered human broncho-epithelial tissue-like assemblies

NASA Technical Reports Server (NTRS)

Goodwin, Thomas J. (Inventor)

2012-01-01

Three-dimensional human broncho-epithelial tissue-like assemblies (TLAs) are produced in a rotating wall vessel (RWV) with microcarriers by coculturing mesenchymal bronchial-tracheal cells (BTC) and bronchial epithelium cells (BEC). These TLAs display structural characteristics and express markers of in vivo respiratory epithelia. TLAs are useful for screening compounds active in lung tissues such as antiviral compounds, cystic fibrosis treatments, allergens, and cytotoxic compounds.

Cellulose factories: advancing bioenergy production from forest trees.

PubMed

Mizrachi, Eshchar; Mansfield, Shawn D; Myburg, Alexander A

2012-04-01

Fast-growing, short-rotation forest trees, such as Populus and Eucalyptus, produce large amounts of cellulose-rich biomass that could be utilized for bioenergy and biopolymer production. Major obstacles need to be overcome before the deployment of these genera as energy crops, including the effective removal of lignin and the subsequent liberation of carbohydrate constituents from wood cell walls. However, significant opportunities exist to both select for and engineer the structure and interaction of cell wall biopolymers, which could afford a means to improve processing and product development. The molecular underpinnings and regulation of cell wall carbohydrate biosynthesis are rapidly being elucidated, and are providing tools to strategically develop and guide the targeted modification required to adapt forest trees for the emerging bioeconomy. Much insight has already been gained from the perturbation of individual genes and pathways, but it is not known to what extent the natural variation in the sequence and expression of these same genes underlies the inherent variation in wood properties of field-grown trees. The integration of data from next-generation genomic technologies applied in natural and experimental populations will enable a systems genetics approach to study cell wall carbohydrate production in trees, and should advance the development of future woody bioenergy and biopolymer crops.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Somerville, Chris

Berkeley Lab's Chris Somerville is a leading authority on the structure and function of plant cell walls, which comprise most of the body mass of higher plants. He views the knowledge of cell wall structure and function as furthering the development of plants with improved usefulness: these plants are strong potential sources of renewable materials and biofuel feedstocks. His scientific expertise defines an ideal match of his interest - in the development of cellulosic and other solar-to-fuel science - with his recent appointment as Director of the Energy Biosciences Institute (EBI). With colleagues in biology, physical sciences, engineering, and environmentalmore » and the social sciences, he now leads the EBI multidisciplinary teams' research efforts to develop next-generation, carbon-neutral transportation fuels.« less

Designer lignins: harnessing the plasticity of lignification

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mottiar, Yaseen; Vanholme, Ruben; Boerjan, Wout

Lignin is a complex polyphenolic constituent of plant secondary cell walls. Inspired largely by the recalcitrance of lignin to biomass processing, plant engineering efforts have routinely sought to alter lignin quantity, composition, and structure by exploiting the inherent plasticity of lignin biosynthesis. More recently, researchers are attempting to strategically design plants for increased degradability by incorporating monomers that lead to a lower degree of polymerisation, reduced hydrophobicity, fewer bonds to other cell wall constituents, or novel chemically labile linkages in the polymer backbone. In addition, the incorporation of value-added structures could help valorise lignin. Designer lignins may satisfy the biologicalmore » requirement for lignification in plants while improving the overall efficiency of biomass utilisation.« less

Designer lignins: harnessing the plasticity of lignification

DOE PAGES

Mottiar, Yaseen; Vanholme, Ruben; Boerjan, Wout; ...

2016-01-15

Lignin is a complex polyphenolic constituent of plant secondary cell walls. Inspired largely by the recalcitrance of lignin to biomass processing, plant engineering efforts have routinely sought to alter lignin quantity, composition, and structure by exploiting the inherent plasticity of lignin biosynthesis. More recently, researchers are attempting to strategically design plants for increased degradability by incorporating monomers that lead to a lower degree of polymerisation, reduced hydrophobicity, fewer bonds to other cell wall constituents, or novel chemically labile linkages in the polymer backbone. In addition, the incorporation of value-added structures could help valorise lignin. Designer lignins may satisfy the biologicalmore » requirement for lignification in plants while improving the overall efficiency of biomass utilisation.« less

Engineering yeast for the expression and secretion of cellulase cocktails

USDA-ARS?s Scientific Manuscript database

Enzyme systems that digest the cellulose in plant cell walls have potential value in the biorefining of renewable feedstocks such as crop residues, straws, and grasses to biofuels and other bioproducts. The bacterium Clostridium cellulovorans is a useful source of biomass-degrading enzymes because ...

Engineering of plants with improved properties as biofuels feedstocks by vessel-specific complementation of xylan biosynthesis mutants

PubMed Central

2012-01-01

Background Cost-efficient generation of second-generation biofuels requires plant biomass that can easily be degraded into sugars and further fermented into fuels. However, lignocellulosic biomass is inherently recalcitrant toward deconstruction technologies due to the abundant lignin and cross-linked hemicelluloses. Furthermore, lignocellulosic biomass has a high content of pentoses, which are more difficult to ferment into fuels than hexoses. Engineered plants with decreased amounts of xylan in their secondary walls have the potential to render plant biomass a more desirable feedstock for biofuel production. Results Xylan is the major non-cellulosic polysaccharide in secondary cell walls, and the xylan deficient irregular xylem (irx) mutants irx7, irx8 and irx9 exhibit severe dwarf growth phenotypes. The main reason for the growth phenotype appears to be xylem vessel collapse and the resulting impaired transport of water and nutrients. We developed a xylan-engineering approach to reintroduce xylan biosynthesis specifically into the xylem vessels in the Arabidopsis irx7, irx8 and irx9 mutant backgrounds by driving the expression of the respective glycosyltransferases with the vessel-specific promoters of the VND6 and VND7 transcription factor genes. The growth phenotype, stem breaking strength, and irx morphology was recovered to varying degrees. Some of the plants even exhibited increased stem strength compared to the wild type. We obtained Arabidopsis plants with up to 23% reduction in xylose levels and 18% reduction in lignin content compared to wild-type plants, while exhibiting wild-type growth patterns and morphology, as well as normal xylem vessels. These plants showed a 42% increase in saccharification yield after hot water pretreatment. The VND7 promoter yielded a more complete complementation of the irx phenotype than the VND6 promoter. Conclusions Spatial and temporal deposition of xylan in the secondary cell wall of Arabidopsis can be manipulated by using the promoter regions of vessel-specific genes to express xylan biosynthetic genes. The expression of xylan specifically in the xylem vessels is sufficient to complement the irx phenotype of xylan deficient mutants, while maintaining low overall amounts of xylan and lignin in the cell wall. This engineering approach has the potential to yield bioenergy crop plants that are more easily deconstructed and fermented into biofuels. PMID:23181474

Endometrial stem cell differentiation into smooth muscle cell: a novel approach for bladder tissue engineering in women.

PubMed

Shoae-Hassani, Alireza; Sharif, Shiva; Seifalian, Alexander M; Mortazavi-Tabatabaei, Seyed Abdolreza; Rezaie, Sassan; Verdi, Javad

2013-10-01

To investigate manufacturing smooth muscle cells (SMCs) for regenerative bladder reconstruction from differentiation of endometrial stem cells (EnSCs), as the recent discovery of EnSCs from the lining of women's uteri, opens up the possibility of using these cells for tissue engineering applications, such as building up natural tissue to repair prolapsed pelvic floors as well as building urinary bladder wall. Human EnSCs that were positive for cluster of differentiation 146 (CD146), CD105 and CD90 were isolated and cultured in Dulbecco's modified Eagle/F12 medium supplemented with myogenic growth factors. The myogenic factors included: transforming growth factor β, platelet-derived growth factor, hepatocyte growth factor and vascular endothelial growth factor. Differentiated SMCs on bioabsorbable polyethylene-glycol and collagen hydrogels were checked for SMC markers by real-time reverse-transcriptase polymerase chain reaction (RT-PCR), western blot (WB) and immunocytochemistry (ICC) analyses. Histology confirmed the growth of SMCs in the hydrogel matrices. The myogenic growth factors decreased the proliferation rate of EnSCs, but they differentiated the human EnSCs into SMCs more efficiently on hydrogel matrices and expressed specific SMC markers including α-smooth muscle actin, desmin, vinculin and calponin in RT-PCR, WB and ICC experiments. The survival rate of cultures on the hydrogel-coated matrices was significantly higher than uncoated cultures. Human EnSCs were successfully differentiated into SMCs, using hydrogels as scaffold. EnSCs may be used for autologous bladder wall regeneration without any immunological complications in women. Currently work is in progress using bioabsorbable nanocomposite materials as EnSC scaffolds for developing urinary bladder wall tissue. © 2013 The Authors. BJU International © 2013 BJU International.

Expanding Imaging Capabilities for Microfluidics: Applicability of Darkfield Internal Reflection Illumination (DIRI) to Observations in Microfluidics

PubMed Central

Kawano, Yoshihiro; Otsuka, Chino; Sanzo, James; Higgins, Christopher; Nirei, Tatsuo; Schilling, Tobias; Ishikawa, Takuji

2015-01-01

Microfluidics is used increasingly for engineering and biomedical applications due to recent advances in microfabrication technologies. Visualization of bubbles, tracer particles, and cells in a microfluidic device is important for designing a device and analyzing results. However, with conventional methods, it is difficult to observe the channel geometry and such particles simultaneously. To overcome this limitation, we developed a Darkfield Internal Reflection Illumination (DIRI) system that improved the drawbacks of a conventional darkfield illuminator. This study was performed to investigate its utility in the field of microfluidics. The results showed that the developed system could clearly visualize both microbubbles and the channel wall by utilizing brightfield and DIRI illumination simultaneously. The methodology is useful not only for static phenomena, such as clogging, but also for dynamic phenomena, such as the detection of bubbles flowing in a channel. The system was also applied to simultaneous fluorescence and DIRI imaging. Fluorescent tracer beads and channel walls were observed clearly, which may be an advantage for future microparticle image velocimetry (μPIV) analysis, especially near a wall. Two types of cell stained with different colors, and the channel wall, can be recognized using the combined confocal and DIRI system. Whole-slide imaging was also conducted successfully using this system. The tiling function significantly expands the observing area of microfluidics. The developed system will be useful for a wide variety of engineering and biomedical applications for the growing field of microfluidics. PMID:25748425

Expanding imaging capabilities for microfluidics: applicability of darkfield internal reflection illumination (DIRI) to observations in microfluidics.

PubMed

Kawano, Yoshihiro; Otsuka, Chino; Sanzo, James; Higgins, Christopher; Nirei, Tatsuo; Schilling, Tobias; Ishikawa, Takuji

2015-01-01

Microfluidics is used increasingly for engineering and biomedical applications due to recent advances in microfabrication technologies. Visualization of bubbles, tracer particles, and cells in a microfluidic device is important for designing a device and analyzing results. However, with conventional methods, it is difficult to observe the channel geometry and such particles simultaneously. To overcome this limitation, we developed a Darkfield Internal Reflection Illumination (DIRI) system that improved the drawbacks of a conventional darkfield illuminator. This study was performed to investigate its utility in the field of microfluidics. The results showed that the developed system could clearly visualize both microbubbles and the channel wall by utilizing brightfield and DIRI illumination simultaneously. The methodology is useful not only for static phenomena, such as clogging, but also for dynamic phenomena, such as the detection of bubbles flowing in a channel. The system was also applied to simultaneous fluorescence and DIRI imaging. Fluorescent tracer beads and channel walls were observed clearly, which may be an advantage for future microparticle image velocimetry (μPIV) analysis, especially near a wall. Two types of cell stained with different colors, and the channel wall, can be recognized using the combined confocal and DIRI system. Whole-slide imaging was also conducted successfully using this system. The tiling function significantly expands the observing area of microfluidics. The developed system will be useful for a wide variety of engineering and biomedical applications for the growing field of microfluidics.

Engineered Tissue–Stent Biocomposites as Tracheal Replacements

PubMed Central

Zhao, Liping; Sundaram, Sumati; Le, Andrew V.; Huang, Angela H.; Zhang, Jiasheng; Hatachi, Go; Beloiartsev, Arkadi; Caty, Michael G.; Yi, Tai; Leiby, Katherine; Gard, Ashley; Kural, Mehmet H.; Gui, Liqiong; Rocco, Kevin A.; Sivarapatna, Amogh; Calle, Elizabeth; Greaney, Allison; Urbani, Luca; Maghsoudlou, Panagiotis; Burns, Alan; DeCoppi, Paolo

2016-01-01

Here we report the creation of a novel tracheal construct in the form of an engineered, acellular tissue–stent biocomposite trachea (TSBT). Allogeneic or xenogeneic smooth muscle cells are cultured on polyglycolic acid polymer–metal stent scaffold leading to the formation of a tissue comprising cells, their deposited collagenous matrix, and the stent material. Thorough decellularization then produces a final acellular tubular construct. Engineered TSBTs were tested as end-to-end tracheal replacements in 11 rats and 3 nonhuman primates. Over a period of 8 weeks, no instances of airway perforation, infection, stent migration, or erosion were observed. Histological analyses reveal that the patent implants remodel adaptively with native host cells, including formation of connective tissue in the tracheal wall and formation of a confluent, columnar epithelium in the graft lumen, although some instances of airway stenosis were observed. Overall, TSBTs resisted collapse and compression that often limit the function of other decellularized tracheal replacements, and additionally do not require any cells from the intended recipient. Such engineered TSBTs represent a model for future efforts in tracheal regeneration. PMID:27520928

Quantification and Control of Wall Effects in Porous Media Experiments

NASA Astrophysics Data System (ADS)

Roth, E. J.; Mays, D. C.; Neupauer, R.; Crimaldi, J. P.

2017-12-01

Fluid flow dynamics in porous media are dominated by media heterogeneity. This heterogeneity can create preferential pathways in which local seepage velocities dwarf system seepage velocities, further complicating an already incomplete understanding of dispersive processes. In physical models of porous media flows, apparatus walls introduce preferential flow paths (i.e., wall effects) that may overwhelm other naturally occurring preferential pathways within the apparatus, leading to deceptive results. We used planar laser-induced fluorescence (PLIF) in conjunction with refractive index matched (RIM) porous media and pore fluid to observe fluid dynamics in the porous media, with particular attention to the region near the apparatus walls in a 17 cm x 8 cm x 7 cm uniform flow cell. Hexagonal close packed spheres were used to create an isotropic, homogenous porous media field in the interior of the apparatus. Visualization of the movement of a fluorescent dye revealed the influence of the wall in creating higher permeability preferential flow paths in an otherwise homogenous media packing. These preferential flow paths extended approximately one half of one sphere diameter from the wall for homogenously packed regions, with a quickly diminishing effect on flow dynamics for homogenous media adjacent to the preferential pathway, but with major influence on flow dynamics for adjoining heterogeneous regions. Multiple approaches to mitigate wall effects were investigated, and a modified wall was created such that the fluid dynamics near the wall mimics the fluid dynamics within the homogenous porous media. This research supports the design of a two-dimensional experimental apparatus that will simulate engineered pumping schemes for use in contaminant remediation. However, this research could benefit the design of fixed bed reactors or other engineering challenges in which vessel walls contribute to unwanted preferential flow.

Evaluation of cell sheet application on one wall bone defect in Macaca nemestrina through periostin expression

NASA Astrophysics Data System (ADS)

Tamin, R. Y.; Soeroso, Y.; Amir, L.; Idrus, E.

2017-08-01

Chronic periodontitis is an oral disease in which the destruction of periodontal tissue leads to tooth loss. Regenerative therapy for attachment cannot be applied to one wall bone defects owing to the minimal existing healthy bone. Tissue engineering in the form of cell sheets has been developed to overcome this limitation. In a previous study, cell sheet application to a one wall bone defect in Macaca nemestrina showed good clinical results. To evaluate the effectiveness of cell sheet application histologically, the level of periostin expression in the gingival crevicular fluid (GCF) of M. nemestrina was determined. Periostin is a 90-kDa protein that regulates coordination and interaction for regeneration and tissue repair. A laboratory observation study was performed to see the differences in periostin levels in samples collected from M. nemestrina’s GCF, where a cell sheet was applied to the bone defect. Gel electrophoresis with SDS-PAGE was performed to detect periostin expression based on its molecular weight and to compare the expression band between the cell sheet and the control at 1, 2, and 3 weeks after treatment. The gel electrophoresis result shows different thicknesses of the protein band around the molecular weight of periostin between the cell sheet groups.

Applications of yeast surface display for protein engineering

PubMed Central

Cherf, Gerald M.; Cochran, Jennifer R.

2015-01-01

The method of displaying recombinant proteins on the surface of Saccharomyces cerevisiae via genetic fusion to an abundant cell wall protein, a technology known as yeast surface display, or simply, yeast display, has become a valuable protein engineering tool for a broad spectrum of biotechnology and biomedical applications. This review focuses on the use of yeast display for engineering protein affinity, stability, and enzymatic activity. Strategies and examples for each protein engineering goal are discussed. Additional applications of yeast display are also briefly presented, including protein epitope mapping, identification of protein-protein interactions, and uses of displayed proteins in industry and medicine. PMID:26060074

A Huygens immersed-finite-element particle-in-cell method for modeling plasma-surface interactions with moving interface

NASA Astrophysics Data System (ADS)

Cao, Huijun; Cao, Yong; Chu, Yuchuan; He, Xiaoming; Lin, Tao

2018-06-01

Surface evolution is an unavoidable issue in engineering plasma applications. In this article an iterative method for modeling plasma-surface interactions with moving interface is proposed and validated. In this method, the plasma dynamics is simulated by an immersed finite element particle-in-cell (IFE-PIC) method, and the surface evolution is modeled by the Huygens wavelet method which is coupled with the iteration of the IFE-PIC method. Numerical experiments, including prototypical engineering applications, such as the erosion of Hall thruster channel wall, are presented to demonstrate features of this Huygens IFE-PIC method for simulating the dynamic plasma-surface interactions.

Thermal energy transformer

NASA Technical Reports Server (NTRS)

Berdahl, C. M.; Thiele, C. L. (Inventor)

1979-01-01

For use in combination with a heat engine, a thermal energy transformer is presented. It is comprised of a flux receiver having a first wall defining therein a radiation absorption cavity for converting solar flux to thermal energy, and a second wall defining an energy transfer wall for the heat engine. There is a heat pipe chamber interposed between the first and second walls having a working fluid disposed within the chamber and a wick lining the chamber for conducting the working fluid from the second wall to the first wall. Thermal energy is transferred from the radiation absorption cavity to the heat engine.

Sugar beet cell wall protein confers fungal and pest resistance in genetically engineered plants

USDA-ARS?s Scientific Manuscript database

Sugar beet biomass and sugar yield are reduced by diseases caused by microbial pathogens and insect pest infestations. Since disease and pest control measures continue to rely on harmful chemical fungicides and insecticides, biotechnological approaches offer an alternate approach for disease and pe...

Hair Follicle-Derived Smooth Muscle Cells and Small Intestinal Submucosa for Engineering Mechanically Robust and Vasoreactive Vascular Media

PubMed Central

Peng, Hao-Fan; Liu, Jin Yu

2011-01-01

Our laboratory recently reported a new source of smooth muscle cells (SMCs) derived from hair follicle (HF) mesenchymal stem cells. HF-SMCs demonstrated high proliferation and clonogenic potential as well as contractile function. In this study, we aimed at engineering the vascular media using HF-SMCs and a natural biomaterial, namely small intestinal submucosa (SIS). Engineering functional vascular constructs required application of mechanical force, resulting in actin reorganization and cellular alignment. In turn, cell alignment was necessary for development of receptor- and nonreceptor-mediated contractility as soon as 24 h after cell seeding. Within 2 weeks in culture, the cells migrated into SIS and secreted collagen and elastin, the two major extracellular matrix components of the vessel wall. At 2 weeks, vascular reactivity increased significantly up to three- to fivefold and mechanical properties were similar to those of native ovine arteries. Taken together, our data demonstrate that the combination of HF-SMCs with SIS resulted in mechanically strong, biologically functional vascular media with potential for arterial implantation. PMID:21083418

LPT. Elevations of low power test building (TAN640 and 641). ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

LPT. Elevations of low power test building (TAN-640 and -641). West and south elevations show stepped shield wall. South and east elevations show pumice block passageway on south side. Reactor cell walls are concrete. One-story parts are pumice block. Metal rollup doors. Ralph M. Parsons 1229-12 ANP/GE-7-640-A-2. November 1956. Approved by INEEL Classification Office for public release. INEEL index code no. 038-0640-00-693-107275 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

Minimally invasive cell-seeded biomaterial systems for injectable/epicardial implantation in ischemic heart disease

PubMed Central

Ravichandran, Rajeswari; Venugopal, Jayarama Reddy; Sundarrajan, Subramanian; Mukherjee, Shayanti; Ramakrishna, Seeram

2012-01-01

Myocardial infarction (MI) is characterized by heart-wall thinning, myocyte slippage, and ventricular dilation. The injury to the heart-wall muscle after MI is permanent, as after an abundant cell loss the myocardial tissue lacks the intrinsic capability to regenerate. New therapeutics are required for functional improvement and regeneration of the infarcted myocardium, to overcome harmful diagnosis of patients with heart failure, and to overcome the shortage of heart donors. In the past few years, myocardial tissue engineering has emerged as a new and ambitious approach for treating MI. Several left ventricular assist devices and epicardial patches have been developed for MI. These devices and acellular/cellular cardiac patches are employed surgically and sutured to the epicardial surface of the heart, limiting the region of therapeutic benefit. An injectable system offers the potential benefit of minimally invasive release into the myocardium either to restore the injured extracellular matrix or to act as a scaffold for cell delivery. Furthermore, intramyocardial injection of biomaterials and cells has opened new opportunities to explore and also to augment the potentials of this technique to ease morbidity and mortality rates owing to heart failure. This review summarizes the growing body of literature in the field of myocardial tissue engineering, where biomaterial injection, with or without simultaneous cellular delivery, has been pursued to enhance functional and structural outcomes following MI. Additionally, this review also provides a complete outlook on the tissue-engineering therapies presently being used for myocardial regeneration, as well as some perceptivity into the possible issues that may hinder its progress in the future. PMID:23271906

Reducing Our Carbon Footprint: Converting Plants to Fuel (LBNL Science at the Theater)

ScienceCinema

Somerville, Chris [Univ. of California, Berkeley, CA (United States)

2018-05-23

Berkeley Lab's Chris Somerville is a leading authority on the structure and function of plant cell walls, which comprise most of the body mass of higher plants. He views the knowledge of cell wall structure and function as furthering the development of plants with improved usefulness: these plants are strong potential sources of renewable materials and biofuel feedstocks. His scientific expertise defines an ideal match of his interest - in the development of cellulosic and other solar-to-fuel science - with his recent appointment as Director of the Energy Biosciences Institute (EBI). With colleagues in biology, physical sciences, engineering, and environmental and the social sciences, he now leads the EBI multidisciplinary teams' research efforts to develop next-generation, carbon-neutral transportation fuels.

Finite element modeling of the cyclic wetting mechanism in the active part of wheat awns.

PubMed

Zickler, Gerald A; Ruffoni, Davide; Dunlop, John W C; Elbaum, Rivka; Weinkamer, Richard; Fratzl, Peter; Antretter, Thomas

2012-12-01

Many plant tissues and organs are capable of moving due to changes in the humidity of the environment, such as the opening of the seed capsule of the ice plant and the opening of the pine cone. These are fascinating examples for the materials engineer, as these tissues are non-living and move solely through the differential swelling of anisotropic tissues and in principle may serve as examples for the bio-inspired design of artificial actuators. In this paper, we model the microstructure of the wild wheat awn (Triticum turgidum ssp. dicoccoides) by finite elements, especially focusing on the specific microscopic features of the active part of the awn. Based on earlier experimental findings, cell walls are modeled as multilayered cylindrical tubes with alternating cellulose fiber orientation in successive layers. It is shown that swelling upon hydration of this system leads to the formation of gaps between the layers, which could act as valves, thus enabling the entry of water into the cell wall. This supports the hypothesis that this plywood-like arrangement of cellulose fibrils enhances the effect of ambient humidity by accelerated water or vapor diffusion along the gaps. The finite element model shows that a certain distribution of axially and tangentially oriented fibers is necessary to generate sufficient tensile stresses within the cell wall to open nanometer-sized gaps between cell wall layers.

Induced mutations in tomato SlExp1 alter cell wall metabolism and delay fruit softening.

PubMed

Minoia, Silvia; Boualem, Adnane; Marcel, Fabien; Troadec, Christelle; Quemener, Bernard; Cellini, Francesco; Petrozza, Angelo; Vigouroux, Jacqueline; Lahaye, Marc; Carriero, Filomena; Bendahmane, Abdelhafid

2016-01-01

Fruit ripening and softening are key traits for many fleshy fruit. Since cell walls play a key role in the softening process, expansins have been investigated to control fruit over ripening and deterioration. In tomato, expression of Expansin 1 gene, SlExp1, during fruit ripening was associated with fruit softening. To engineer tomato plants with long shelf life, we screened for mutant plants impaired in SlExp1 function. Characterization of two induced mutations, Slexp1-6_W211S, and Slexp1-7_Q213Stop, showed that SlExp1 loss of function leads to enhanced fruit firmness and delayed fruit ripening. Analysis of cell wall polysaccharide composition of Slexp1-7_Q213Stop mutant pointed out significant differences for uronic acid, neutral sugar and total sugar contents. Hemicelluloses chemistry analysis by endo-β-1,4-d-glucanase hydrolysis and MALDI-TOF spectrometry revealed that xyloglucan structures were affected in the fruit pericarp of Slexp1-7_Q213Stop mutant. Altogether, these results demonstrated that SlExp1 loss of function mutants yield firmer and late ripening fruits through modification of hemicellulose structure. These SlExp1 mutants represent good tools for breeding long shelf life tomato lines with contrasted fruit texture as well as for the understanding of the cell wall polysaccharide assembly dynamics in fleshy fruits. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

CONSTRUCTION PROGRESS PHOTO OF HOT PILOT PLANT (CPP640) LOOKING NORTHWEST, ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

CONSTRUCTION PROGRESS PHOTO OF HOT PILOT PLANT (CPP-640) LOOKING NORTHWEST, SHOWING FORMING FOR NORTH WALLS OF CELLS 1, 4 AND 5; CONSTRUCTION 21 PERCENT COMPLETE. INL PHOTO NUMBER NRTS-60-1874. Holmes, Photographer, 4/21/1960 - Idaho National Engineering Laboratory, Idaho Chemical Processing Plant, Fuel Reprocessing Complex, Scoville, Butte County, ID

Lipid engineering reveals regulatory roles for membrane fluidity in yeast flocculation and oxygen-limited growth.

PubMed

Degreif, Daniel; de Rond, Tristan; Bertl, Adam; Keasling, Jay D; Budin, Itay

2017-05-01

Cells modulate lipid metabolism in order to maintain membrane homeostasis. Here we use a metabolic engineering approach to manipulate the stoichiometry of fatty acid unsaturation, a regulator of cell membrane fluidity, in Saccharomyces cerevisiae. Unexpectedly, reduced lipid unsaturation triggered cell-cell adhesion (flocculation), a phenomenon characteristic of industrial yeast but uncommon in laboratory strains. We find that ER lipid saturation sensors induce expression of FLO1 - encoding a cell wall polysaccharide binding protein - independently of its canonical regulator. In wild-type cells, Flo1p-dependent flocculation occurs under oxygen-limited growth, which reduces unsaturated lipid synthesis and thus serves as the environmental trigger for flocculation. Transcriptional analysis shows that FLO1 is one of the most highly induced genes in response to changes in lipid unsaturation, and that the set of membrane fluidity-sensitive genes is globally activated as part of the cell's long-term response to hypoxia during fermentation. Our results show how the lipid homeostasis machinery of budding yeast is adapted to carry out a broad response to an environmental stimulus important in biotechnology. Copyright © 2017 International Metabolic Engineering Society. All rights reserved.

Process for forming a long gas turbine engine blade having a main wall with a thin portion near a tip

DOEpatents

Campbell, Christian X; Thomaidis, Dimitrios

2014-05-13

A process is provided for forming an airfoil for a gas turbine engine involving: forming a casting of a gas turbine engine airfoil having a main wall and an interior cavity, the main wall having a wall thickness extending from an external surface of the outer wall to the interior cavity, an outer section of the main wall extending from a location between a base and a tip of the airfoil casting to the tip having a wall thickness greater than a final thickness. The process may further involve effecting movement, using a computer system, of a material removal apparatus and the casting relative to one another such that a layer of material is removed from the casting at one or more radial portions along the main wall of the casting.

Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response

PubMed Central

Tsai, Alex Yi-Lin; Chan, Kin; Ho, Chi-Yip; Canam, Thomas; Capron, Resmi; Master, Emma R.; Bräutigam, Katharina

2017-01-01

The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to the hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that transgenic expression of microbial cell wall modifying enzymes induces transcriptome responses similar to those observed in defense responses. PMID:28253318

Dynamics of biomass partitioning, stem gene expression, cell wall biosynthesis, and sucrose accumulation during development of Sorghum bicolor.

PubMed

McKinley, Brian; Rooney, William; Wilkerson, Curtis; Mullet, John

2016-11-01

Biomass accumulated preferentially in leaves of the sweet sorghum Della until floral initiation, then stems until anthesis, followed by panicles until grain maturity, and apical tillers. Sorghum stem RNA-seq transcriptome profiles and composition data were collected for approximately 100 days of development beginning at floral initiation. The analysis identified >200 differentially expressed genes involved in stem growth, cell wall biology, and sucrose accumulation. Genes encoding expansins and xyloglucan endotransglucosylase/hydrolases were differentially expressed in growing stem internodes. Genes encoding enzymes involved in the synthesis of cellulose, lignin, and glucuronoarabinoxylan were expressed at elevated levels in stems until approximately 7 days before anthesis and then down-regulated. CESA genes involved in primary and secondary cell wall synthesis showed different temporal patterns of expression. Following floral initiation, the level of sucrose and other non-structural carbohydrates increased to approximately 50% of the stem's dry weight. Stem sucrose accumulation was inversely correlated with >100-fold down-regulation of SbVIN1, a gene encoding a vacuolar invertase. Accumulation of stem sucrose was also correlated with cessation of leaf and stem growth at anthesis, decreased expression of genes involved in stem cell wall synthesis, and approximately 10-fold lower expression of SbSUS4, a gene encoding sucrose synthase that generates UDP-glucose from sucrose for cell wall biosynthesis. Genes for mixed linkage glucan synthesis (CSLF) and turnover were expressed at high levels in stems throughout development. Overall, the stem transcription profile resource and the genes and regulatory dynamics identified in this study will be useful for engineering sorghum stem composition for improved conversion to biofuels and bio-products. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

An update to space biomedical research: tissue engineering in microgravity bioreactors.

PubMed

Barzegari, Abolfazl; Saei, Amir Ata

2012-01-01

The severe need for constructing replacement tissues in organ transplanta-tion has necessitated the development of tissue engineering approaches and bioreactors that can bring these approaches to reality. The inherent limitations of conventional bioreactors in generating realistic tissue constructs led to the devise of the microgravity tissue engineering that uses Rotating Wall Vessel (RWV) bioreactors initially developed by NASA. In this review article, we intend to highlight some major advances and accomplishments in the rapidly-growing field of tissue engineering that could not be achieved without using microgravity. Research is now focused on assembly of 3 dimensional (3D) tissue fragments from various cell types in human body such as chon-drocytes, osteoblasts, embryonic and mesenchymal stem cells, hepatocytes and pancreas islet cells. Hepatocytes cultured under microgravity are now being used in extracorporeal bioartificial liver devices. Tissue constructs can be used not only in organ replacement therapy, but also in pharmaco-toxicology and food safety assessment. 3D models of vari-ous cancers may be used in studying cancer development and biology or in high-throughput screening of anticancer drug candidates. Finally, 3D heterogeneous assemblies from cancer/immune cells provide models for immunotherapy of cancer. Tissue engineering in (simulated) microgravity has been one of the stunning impacts of space research on biomedical sciences and their applications on earth.

Engineering secondary cell wall deposition in plants

PubMed Central

Yang, Fan; Mitra, Prajakta; Zhang, Ling; Prak, Lina; Verhertbruggen, Yves; Kim, Jin-Sun; Sun, Lan; Zheng, Kejian; Tang, Kexuan; Auer, Manfred; Scheller, Henrik V; Loqué, Dominique

2013-01-01

Lignocellulosic biomass was used for thousands of years as animal feed and is now considered a great sugar source for biofuels production. It is composed mostly of secondary cell walls built with polysaccharide polymers that are embedded in lignin to reinforce the cell wall structure and maintain its integrity. Lignin is the primary material responsible for biomass recalcitrance to enzymatic hydrolysis. During plant development, deep reductions of lignin cause growth defects and often correlate with the loss of vessel integrity that adversely affects water and nutrient transport in plants. The work presented here describes a new approach to decrease lignin content while preventing vessel collapse and introduces a new strategy to boost transcription factor expression in native tissues. We used synthetic biology tools in Arabidopsis to rewire the secondary cell network by changing promoter-coding sequence associations. The result was a reduction in lignin and an increase in polysaccharide depositions in fibre cells. The promoter of a key lignin gene, C4H, was replaced by the vessel-specific promoter of transcription factor VND6. This rewired lignin biosynthesis specifically for vessel formation while disconnecting C4H expression from the fibre regulatory network. Secondly, the promoter of the IRX8 gene, secondary cell wall glycosyltransferase, was used to express a new copy of the fibre transcription factor NST1, and as the IRX8 promoter is induced by NST1, this also created an artificial positive feedback loop (APFL). The combination of strategies—lignin rewiring with APFL insertion—enhances polysaccharide deposition in stems without over-lignifying them, resulting in higher sugar yields after enzymatic hydrolysis. PMID:23140549

Engine Research Building’s Central Control Room

NASA Image and Video Library

1948-07-21

Operators in the Engine Research Building’s Central Control Room at the National Advisory Committee for Aeronautics (NACA) Lewis Flight Propulsion Laboratory. The massive 4.25-acre Engine Research Building contains dozens of test cells, test stands, and altitude chambers. A powerful collection of compressors and exhausters located in the central portion of the basement provided process air and exhaust for these test areas. This system is connected to similar process air systems in the laboratory’s other large test facilities. The Central Control Room coordinates this activity and communicates with the local utilities. This photograph was taken just after a major upgrade to the control room in 1948. The panels on the wall contain rudimentary floor plans of the different Engine Research Building sections with indicator lights and instrumentation for each test cell. The process air equipment included 12 exhausters, four compressors, a refrigeration system, cooling water, and an exhaust system. The operators in the control room kept in contact with engineers running the process air system and those conducting the tests in the test cells. The operators also coordinated with the local power companies to make sure enough electricity was available to operate the powerful compressors and exhausters.

Bacteriophage endolysins as novel antimicrobials

PubMed Central

Schmelcher, Mathias; Donovan, David M; Loessner, Martin J

2013-01-01

Endolysins are enzymes used by bacteriophages at the end of their replication cycle to degrade the peptidoglycan of the bacterial host from within, resulting in cell lysis and release of progeny virions. Due to the absence of an outer membrane in the Gram-positive bacterial cell wall, endolysins can access the peptidoglycan and destroy these organisms when applied externally, making them interesting antimicrobial candidates, particularly in light of increasing bacterial drug resistance. This article reviews the modular structure of these enzymes, in which cell wall binding and catalytic functions are separated, as well as their mechanism of action, lytic activity and potential as antimicrobials. It particularly focuses on molecular engineering as a means of optimizing endolysins for specific applications, highlights new developments that may render these proteins active against Gram-negative and intracellular pathogens and summarizes the most recent applications of endolysins in the fields of medicine, food safety, agriculture and biotechnology. PMID:23030422

Critical review on the mechanisms of maturation stress generation in trees

PubMed Central

Clair, Bruno

2016-01-01

Trees control their posture by generating asymmetric mechanical stress around the periphery of the trunk or branches. This stress is produced in wood during the maturation of the cell wall. When the need for reaction is high, it is accompanied by strong changes in cell organization and composition called reaction wood, namely compression wood in gymnosperms and tension wood in angiosperms. The process by which stress is generated in the cell wall during its formation is not yet known, and various hypothetical mechanisms have been proposed in the literature. Here we aim at discriminating between these models. First, we summarize current knowledge about reaction wood structure, state and behaviour relevant to the understanding of maturation stress generation. Then, the mechanisms proposed in the literature are listed and discussed in order to identify which can be rejected based on their inconsistency with current knowledge at the frontier between plant science and mechanical engineering. PMID:27605169

Translating textiles to tissue engineering: Creation and evaluation of microporous, biocompatible, degradable scaffolds using industry relevant manufacturing approaches and human adipose derived stem cells.

PubMed

Haslauer, Carla M; Avery, Matthew R; Pourdeyhimi, Behnam; Loboa, Elizabeth G

2015-07-01

Polymeric scaffolds have emerged as a means of generating three-dimensional tissues, such as for the treatment of bone injuries and nonunions. In this study, a fibrous scaffold was designed using the biocompatible, degradable polymer poly-lactic acid in combination with a water dispersible sacrificial polymer, EastONE. Fibers were generated via industry relevant, facile scale-up melt-spinning techniques with an islands-in-the-sea geometry. Following removal of EastONE, a highly porous fiber remained possessing 12 longitudinal channels and pores throughout all internal and external fiber walls. Weight loss and surface area characterization confirmed the generation of highly porous fibers as observed via focused ion beam/scanning electron microscopy. Porous fibers were then knit into a three-dimensional scaffold and seeded with human adipose-derived stem cells (hASC). Confocal microscopy images confirmed hASC attachment to the fiber walls and proliferation throughout the knit structure. Quantification of cell-mediated calcium accretion following culture in osteogenic differentiation medium confirmed hASC differentiation throughout the porous constructs. These results suggest incorporation of a sacrificial polymer within islands-in-the-sea fibers generates a highly porous scaffold capable of supporting stem cell viability and differentiation with the potential to generate large three-dimensional constructs for bone regeneration and/or other tissue engineering applications. © 2014 Wiley Periodicals, Inc.

Engine room, showing engine generator foundation and inlet louvers, looking ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

Engine room, showing engine generator foundation and inlet louvers, looking southwest. Note fuel gauge on west wall, left of fuse box, and exhaust pipe through south wall. - Western Union Telegraph Company, Jennerstown Relay, Laurel Summit Road off U.S. 30, Laughlintown, Westmoreland County, PA

Microbial lipid extraction from Lipomyces starkeyi using irreversible electroporation.

PubMed

Karim, Ahasanul; Yousuf, Abu; Islam, M Amirul; Naif, Yasir H; Faizal, Che Ku Mohammad; Alam, Md Zahangir; Pirozzi, Domenico

2018-02-21

The aim of the study was to investigate the feasibility of using irreversible electroporation (EP) as a microbial cell disruption technique to extract intracellular lipid within short time and in an eco-friendly manner. An EP circuit was designed and fabricated to obtain 4 kV with frequency of 100 Hz of square waves. The yeast cells of Lipomyces starkeyi (L. starkeyi) were treated by EP for 2-10 min where the distance between electrodes was maintained at 2, 4, and 6 cm. Colony forming units (CFU) were counted to observe the cell viability under the high voltage electric field. The forces of the pulsing electric field caused significant damage to the cell wall of L. starkeyi and the disruption of microbial cells was visualized by field emission scanning electron microscopic (FESEM) image. After breaking the cell wall, lipid was extracted and measured to assess the efficiency of EP over other techniques. The extent of cell inactivation was up to 95% when the electrodes were placed at the distance of 2 cm, which provided high treatment intensity (36.7 kWh m -3 ). At this condition, maximum lipid (63 mg g -1 ) was extracted when the biomass was treated for 10 min. During the comparison, EP could extract 31.88% lipid while the amount was 11.89% for ultrasonic and 16.8% for Fenton's reagent. The results recommend that the EP is a promising technique for lowering the time and solvent usage for lipid extraction from microbial biomass. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 2018. © 2018 American Institute of Chemical Engineers.

Utilization of non-conventional systems for conversion of biomass to food components: Potential for utilization of algae in engineered foods

NASA Technical Reports Server (NTRS)

Karel, M.; Kamarei, A. R.; Nakhost, Z.

1985-01-01

The major nutritional components of the green algae (Scenedesmus obliquus) grown in a Constant Cell density Apparatus were determined. Suitable methodology to prepare proteins from which three major undesirable components of these cells (i.e., cell walls, nucleic acids, and pigments) were either removed or substantially reduced was developed. Results showed that processing of green algae to protein isolate enhances its potential nutritional and organoleptic acceptability as a diet component in a Controlled Ecological Life Support System.

Application of Freeze-Dried Powders of Genetically Engineered Microbial Strains as Adsorbents for Rare Earth Metal Ions.

PubMed

Moriwaki, Hiroshi; Masuda, Reiko; Yamazaki, Yuki; Horiuchi, Kaoru; Miyashita, Mari; Kasahara, Jun; Tanaka, Tatsuhito; Yamamoto, Hiroki

2016-10-12

The adsorption behaviors of the rare earth metal ions onto freeze-dried powders of genetically engineered microbial strains were compared. Cell powders obtained from four kinds of strains, Bacillus subtilis 168 wild type (WT), lipoteichoic acid-defective (ΔLTA), wall teichoic acid-defective (ΔWTA), and cell wall hydrolases-defective (EFKYOJLp) strains, were used as an adsorbent of the rare earth metal ions at pH 3. The adsorption ability of the rare earth metal ions was in the order of EFKYOJLp > WT > ΔLTA > ΔWTA. The order was the same as the order of the phosphorus quantity of the strains. This result indicates that the main adsorption sites for the ions are the phosphate groups and the teichoic acids, LTA and WTA, that contribute to the adsorption of the rare earth metal ions onto the cell walls. The contribution of WTA was clearly greater than that of LTA. Each microbial powder was added to a solution containing 16 kinds of rare earth metal ions, and the removals (%) of each rare earth metal ion were obtained. The scandium ion showed the highest removal (%), while that of the lanthanum ion was the lowest for all the microbial powders. Differences in the distribution coefficients between the kinds of lanthanide ions by the EFKYOJLp and ΔWTA powders were greater than those of the other strains. Therefore, the EFKYOJLp and ΔWTA powders could be applicable for the selective extraction of the lanthanide ions. The ΔLTA powder coagulated by mixing with a rare earth metal ion, although no sedimentation of the WT or ΔWTA powder with a rare earth metal ion was observed under the same conditions. The EFKYOJLp powder was also coagulated, but its flocculating activity was lower than that of ΔLTA. The ΔLTA and EFKYOJLp powders have a long shape compared to those of the WT or ΔWTA strain. The shapes of the cells will play an important role in the sedimentation of the microbial powders with rare earth metal ions. As the results, three kinds of the genetically engineered microbial powders revealed unique adsorption behaviors of the rare earth metal ions.

Ceramic gas turbine shroud

DOEpatents

Shi, Jun; Green, Kevin E.

2014-07-22

An example gas turbine engine shroud includes a first annular ceramic wall having an inner side for resisting high temperature turbine engine gasses and an outer side with a plurality of radial slots. A second annular metallic wall is positioned radially outwardly of and enclosing the first annular ceramic wall and has a plurality of tabs in communication with the slot of the first annular ceramic wall. The tabs of the second annular metallic wall and slots of the first annular ceramic wall are in communication such that the first annular ceramic wall and second annular metallic wall are affixed.

Optimized suspension culture: the rotating-wall vessel

NASA Technical Reports Server (NTRS)

Hammond, T. G.; Hammond, J. M.

2001-01-01

Suspension culture remains a popular modality, which manipulates mechanical culture conditions to maintain the specialized features of cultured cells. The rotating-wall vessel is a suspension culture vessel optimized to produce laminar flow and minimize the mechanical stresses on cell aggregates in culture. This review summarizes the engineering principles, which allow optimal suspension culture conditions to be established, and the boundary conditions, which limit this process. We suggest that to minimize mechanical damage and optimize differentiation of cultured cells, suspension culture should be performed in a solid-body rotation Couette-flow, zero-headspace culture vessel such as the rotating-wall vessel. This provides fluid dynamic operating principles characterized by 1) solid body rotation about a horizontal axis, characterized by colocalization of cells and aggregates of different sedimentation rates, optimally reduced fluid shear and turbulence, and three-dimensional spatial freedom; and 2) oxygenation by diffusion. Optimization of suspension culture is achieved by applying three tradeoffs. First, terminal velocity should be minimized by choosing microcarrier beads and culture media as close in density as possible. Next, rotation in the rotating-wall vessel induces both Coriolis and centrifugal forces, directly dependent on terminal velocity and minimized as terminal velocity is minimized. Last, mass transport of nutrients to a cell in suspension culture depends on both terminal velocity and diffusion of nutrients. In the transduction of mechanical culture conditions into cellular effects, several lines of evidence support a role for multiple molecular mechanisms. These include effects of shear stress, changes in cell cycle and cell death pathways, and upstream regulation of secondary messengers such as protein kinase C. The discipline of suspension culture needs a systematic analysis of the relationship between mechanical culture conditions and biological effects, emphasizing cellular processes important for the industrial production of biological pharmaceuticals and devices.

New insight into artifactual phenomena during in vitro toxicity assessment of engineered nanoparticles: study of TNF-α adsorption on alumina oxide nanoparticle.

PubMed

Pailleux, Mélanie; Boudard, Delphine; Pourchez, Jérémie; Forest, Valérie; Grosseau, Philippe; Cottier, Michèle

2013-04-01

Biomolecules can be adsorbed on nanoparticles (NPs) and degraded during in vitro toxicity assays. These artifactual phenomena could lead to misinterpretation of biological activity, such as false-negative results. To avoid possible underestimation of cytokine release after contact between NP and cells, we propose a methodology to account for these artifactual phenomena and lead to accurate measurements. We focused on the pro-inflammatory cytokine tumor necrosis factor TNF-α. We studied well-characterized boehmite engineered NP [aluminum oxide hydroxide, AlO(OH)]. The rate of TNF-α degradation and its adsorption (on boehmite and on the walls of wells) were determined in cell-free conditions by adding a known TNF-α concentration (1500 pg/ml) under various experimental conditions. After a 24-h incubation, we quantified that 7 wt.% of the initial TNF-α was degraded over time, 6 wt.% adsorbed on the walls of 96-well plates, and 13 wt.% adsorbed on the boehmite surface. Finally, boehmite NP were incubated with murine macrophages (RAW 264.7 cell line). The release of TNF-α was assessed for boehmite NP and the experimental data were corrected considering the artifactual phenomena, which accounted for about 20-30% of the total. Copyright © 2013 Elsevier Ltd. All rights reserved.

Systems Level Engineering of Plant Cell Wall Biosynthesis to Improve Biofuel Feedstock Quality

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hazen, Samuel

2013-09-27

Our new regulatory model of cell wall biosynthesis proposes original network architecture with several newly incorporated components. The mapped set of protein-DNA interactions will serve as a foundation for 1) understanding the regulation of a complex and integral plant component and 2) the manipulation of crop species for biofuel and biotechnology purposes. This study revealed interesting and novel aspects of grass growth and development and further enforce the importance of a grass model system. By functionally characterizing a suite of genes, we have begun to improve the sparse model for transcription regulation of biomass accumulation in grasses. In the process,more » we have advanced methodology and brachy molecular genetic tools that will serve as valuable community resource.« less

Multi-faceted Approach to Vaccine Development Against Escherichia coli O157:H7

DTIC Science & Technology

2006-03-15

36 Intimin as a vaccine candidate .................................................................................. 39 Transgenic Plant -based...41 Transgenic Plant -based Vaccines Unique advantages of plant vaccines Transgenic plants have been engineered to produce recombinant...strategies. Foremost, the vaccine can be delivered by ingestion of the edible part of the transgenic plant . The plant cell wall acts as a capsule that

LPT. Low power assembly and test building (TAN640). Camera facing ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

LPT. Low power assembly and test building (TAN-640). Camera facing west. Rollup doors to each test cell face east. Concrete walls poured in place. Apparatus at right of view was part of a post-ANP program. INEEL negative no. HD-40-1-1 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

Charge-based separation of particles and cells with similar sizes via the wall-induced electrical lift.

PubMed

Thomas, Cory; Lu, Xinyu; Todd, Andrew; Raval, Yash; Tzeng, Tzuen-Rong; Song, Yongxin; Wang, Junsheng; Li, Dongqing; Xuan, Xiangchun

2017-01-01

The separation of particles and cells in a uniform mixture has been extensively studied as a necessity in many chemical and biomedical engineering and research fields. This work demonstrates a continuous charge-based separation of fluorescent and plain spherical polystyrene particles with comparable sizes in a ψ-shaped microchannel via the wall-induced electrical lift. The effects of both the direct current electric field in the main-branch and the electric field ratio in between the inlet branches for sheath fluid and particle mixture are investigated on this electrokinetic particle separation. A Lagrangian tracking method based theoretical model is also developed to understand the particle transport in the microchannel and simulate the parametric effects on particle separation. Moreover, the demonstrated charge-based separation is applied to a mixture of yeast cells and polystyrene particles with similar sizes. Good separation efficiency and purity are achieved for both the cells and the particles. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Three-dimensional wet-electrospun poly(lactic acid)/multi-wall carbon nanotubes scaffold induces differentiation of human menstrual blood-derived stem cells into germ-like cells.

PubMed

Eyni, Hossein; Ghorbani, Sadegh; Shirazi, Reza; Salari Asl, Leila; P Beiranvand, Shahram; Soleimani, Masoud

2017-09-01

Infertility caused by the disruption or absence of germ cells is a major and largely incurable medical problem. Germ cells (i.e., sperm or egg) play a key role in the transmission of genetic and epigenetic information across generations. Generation of gametes derived in vitro from stem cells hold promising prospects which could potentially help infertile men and women. Menstrual blood-derived stem cells are a unique stem cell source. Evidence suggests that menstrual blood-derived stem cells exhibit a multi-lineage potential and have attracted extensive attention in regenerative medicine. To maintain the three-dimensional structure of natural extra cellular matrices in vitro, scaffolds can do this favor and mimic a microenvironment for cell proliferation and differentiation. According to previous studies, poly(lactic acid) and multi-wall carbon nanotubes have been introduced as novel and promising biomaterials for the proliferation and differentiation of stem cells. Some cell types have been successfully grown on a matrix containing carbon nanotubes in tissue engineering but there is no report for this material to support stem cells differentiation into germ cells lineage. This study designed a 3D wet-electrospun poly(lactic acid) and poly(lactic acid)/multi-wall carbon nanotubes composite scaffold to compare infiltration, proliferation, and differentiation potential of menstrual blood-derived stem cells toward germ cell lineage with 2D culture. Our primary data revealed that the fabricated scaffold has mechanical and biological suitable qualities for supporting and attachments of stem cells. The differentiated menstrual blood-derived stem cells tracking in scaffolds using scanning electron microscopy confirmed cell attachment, aggregation, and distribution on the porous scaffold. Based on the differentiation assay by RT-PCR analysis, stem cells and germ-like cells markers were expressed in 3D groups as well as 2D one. It seems that poly(lactic acid)/multi-wall carbon nanotubes scaffold-seeded menstrual blood-derived stem cells could be viewed as a novel, safe, and accessible construct for these cells, as they enhance germ-like generation from menstrual blood-derived stem cells.

The impact of simulated and real microgravity on bone cells and mesenchymal stem cells.

PubMed

Ulbrich, Claudia; Wehland, Markus; Pietsch, Jessica; Aleshcheva, Ganna; Wise, Petra; van Loon, Jack; Magnusson, Nils; Infanger, Manfred; Grosse, Jirka; Eilles, Christoph; Sundaresan, Alamelu; Grimm, Daniela

2014-01-01

How microgravity affects the biology of human cells and the formation of 3D cell cultures in real and simulated microgravity (r- and s-µg) is currently a hot topic in biomedicine. In r- and s-µg, various cell types were found to form 3D structures. This review will focus on the current knowledge of tissue engineering in space and on Earth using systems such as the random positioning machine (RPM), the 2D-clinostat, or the NASA-developed rotating wall vessel bioreactor (RWV) to create tissue from bone, tumor, and mesenchymal stem cells. To understand the development of 3D structures, in vitro experiments using s-µg devices can provide valuable information about modulations in signal-transduction, cell adhesion, or extracellular matrix induced by altered gravity conditions. These systems also facilitate the analysis of the impact of growth factors, hormones, or drugs on these tissue-like constructs. Progress has been made in bone tissue engineering using the RWV, and multicellular tumor spheroids (MCTS), formed in both r- and s-µg, have been reported and were analyzed in depth. Currently, these MCTS are available for drug testing and proteomic investigations. This review provides an overview of the influence of µg on the aforementioned cells and an outlook for future perspectives in tissue engineering.

The Impact of Simulated and Real Microgravity on Bone Cells and Mesenchymal Stem Cells

PubMed Central

Wehland, Markus; Pietsch, Jessica; Aleshcheva, Ganna; Wise, Petra; van Loon, Jack; Magnusson, Nils; Infanger, Manfred; Grosse, Jirka; Eilles, Christoph

2014-01-01

How microgravity affects the biology of human cells and the formation of 3D cell cultures in real and simulated microgravity (r- and s-µg) is currently a hot topic in biomedicine. In r- and s-µg, various cell types were found to form 3D structures. This review will focus on the current knowledge of tissue engineering in space and on Earth using systems such as the random positioning machine (RPM), the 2D-clinostat, or the NASA-developed rotating wall vessel bioreactor (RWV) to create tissue from bone, tumor, and mesenchymal stem cells. To understand the development of 3D structures, in vitro experiments using s-µg devices can provide valuable information about modulations in signal-transduction, cell adhesion, or extracellular matrix induced by altered gravity conditions. These systems also facilitate the analysis of the impact of growth factors, hormones, or drugs on these tissue-like constructs. Progress has been made in bone tissue engineering using the RWV, and multicellular tumor spheroids (MCTS), formed in both r- and s-µg, have been reported and were analyzed in depth. Currently, these MCTS are available for drug testing and proteomic investigations. This review provides an overview of the influence of µg on the aforementioned cells and an outlook for future perspectives in tissue engineering. PMID:25110709

ZmNST3 and ZmNST4 are master switches for secondary wall deposition in maize (Zea mays L.).

PubMed

Xiao, Wenhan; Yang, Yue; Yu, Jingjuan

2018-01-01

Secondary walls are the most abundant biomass produced by plants, and they consist mainly of lignin, cellulose and hemicellulose. Understanding how secondary wall biosynthesis is regulated could potentially provide genetic tools for engineering biomass components, especially in maize and Sorghum bicolor. Although many works have focused on secondary wall biosynthesis in dicotyledons, little has been reported for these monocotyledons. In this study, we cloned two NAC transcriptional factor genes, ZmNST3 and ZmNST4, and analyzed their functions in maize secondary wall formation process. ZmNST3 and ZmNST4 were expressed specifically in secondary wall-forming cells, expression of ZmNST3/4 can restore the pendent phenotype of Arabidopsis nst1nst3 double mutant. ZmNST3/4-overexpressing Arabidopsis and maize displayed a thickened secondary wall in the stem, and knockdown maize showed defective secondary wall deposition. ZmNST3/4 could regulate the expression of ZmMYB109/128/149. Our results revealed that ZmNST3/4 are master switches of the maize secondary wall biosynthesis process and provides new evidence that the secondary wall regulatory pathway is conserved in different plant species. Copyright © 2017. Published by Elsevier B.V.

Functional analyses of cellulose synthase genes in flax (Linum usitatissimum) by virus-induced gene silencing.

PubMed

Chantreau, Maxime; Chabbert, Brigitte; Billiard, Sylvain; Hawkins, Simon; Neutelings, Godfrey

2015-12-01

Flax (Linum usitatissimum) bast fibres are located in the stem cortex where they play an important role in mechanical support. They contain high amounts of cellulose and so are used for linen textiles and in the composite industry. In this study, we screened the annotated flax genome and identified 14 distinct cellulose synthase (CESA) genes using orthologous sequences previously identified. Transcriptomics of 'primary cell wall' and 'secondary cell wall' flax CESA genes showed that some were preferentially expressed in different organs and stem tissues providing clues as to their biological role(s) in planta. The development for the first time in flax of a virus-induced gene silencing (VIGS) approach was used to functionally evaluate the biological role of different CESA genes in stem tissues. Quantification of transcript accumulation showed that in many cases, silencing not only affected targeted CESA clades, but also had an impact on other CESA genes. Whatever the targeted clade, inactivation by VIGS affected plant growth. In contrast, only clade 1- and clade 6-targeted plants showed modifications in outer-stem tissue organization and secondary cell wall formation. In these plants, bast fibre number and structure were severely impacted, suggesting that the targeted genes may play an important role in the establishment of the fibre cell wall. Our results provide new fundamental information about cellulose biosynthesis in flax that should facilitate future plant improvement/engineering. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Application of Lactobacillus johnsonii expressing phage endolysin for control of Clostridium perfringens.

PubMed

Gervasi, T; Lo Curto, R; Minniti, E; Narbad, A; Mayer, M J

2014-10-01

Clostridium perfringens is frequently found in food and the environment and produces potent toxins that have a negative impact on both human and animal health and particularly on the poultry industry. Lactobacillus johnsonii FI9785, isolated from the chicken gastrointestinal tract, has been demonstrated to exclude Cl. perfringens in poultry. We have investigated the interaction of wild-type Lact. johnsonii FI9785 or an engineered strain expressing a cell wall-hydrolysing endolysin with Cl. perfringens in vitro, using a batch culture designed to simulate human gastrointestinal tract conditions. Co-culture experiments indicated that acid production by Lact. johnsonii is important in pathogen control. The co-culture of the endolysin-secreting Lact. johnsonii with Cl. perfringens showed that the engineered strain had the potential to control the pathogen, but the ability to reduce Cl. perfringens numbers was not consistent. Results obtained indicate that survival of high numbers of Lact. johnsonii will be essential for effective pathogen control. Significance and impact of the study: The bacterium Lactobacillus johnsonii FI9785 reduces numbers of the pathogen Clostridium perfringens in vitro. Biocontrol was improved by engineering the strain to produce and export a cell wall-hydrolysing endolysin, but good survival of the producer strain is essential. The production of bacteriophage endolysins by commensal bacteria has the potential to improve competitive exclusion of pathogens in the gastrointestinal tract. © 2014 The Society for Applied Microbiology.

A Dual-Mode Bioreactor System for Tissue Engineered Vascular Models.

PubMed

Bono, N; Meghezi, S; Soncini, M; Piola, M; Mantovani, D; Fiore, Gianfranco Beniamino

2017-06-01

In the past decades, vascular tissue engineering has made great strides towards bringing engineered vascular tissues to the clinics and, in parallel, obtaining in-lab tools for basic research. Herein, we propose the design of a novel dual-mode bioreactor, useful for the fabrication (construct mode) and in vitro stimulation (culture mode) of collagen-based tubular constructs. Collagen-based gels laden with smooth muscle cells (SMCs) were molded directly within the bioreactor culture chamber. Based on a systematic characterization of the bioreactor culture mode, constructs were subjected to 10% cyclic strain at 0.5 Hz for 5 days. The effects of cyclic stimulation on matrix re-arrangement and biomechanical/viscoelastic properties were examined and compared vs. statically cultured constructs. A thorough comparison of cell response in terms of cell localization and expression of contractile phenotypic markers was carried out as well. We found that cyclic stimulation promoted cell-driven collagen matrix bi-axial compaction, enhancing the mechanical strength of strained samples with respect to static controls. Moreover, cyclic strain positively affected SMC behavior: cells maintained their contractile phenotype and spread uniformly throughout the whole wall thickness. Conversely, static culture induced a noticeable polarization of cell distribution to the outer rim of the constructs and a sharp reduction in total cell density. Overall, coupling the use of a novel dual-mode bioreactor with engineered collagen-gel-based tubular constructs demonstrated to be an interesting technology to investigate the modulation of cell and tissue behavior under controlled mechanically conditioned in vitro maturation.

IET. Movable test cell building (TAN624). Plans, sections, and elevations ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

IET. Movable test cell building (TAN-624). Plans, sections, and elevations show trapezoidal shape of front/rear elevations, vertical sliding door panels, wheels, periscope and camera locations, fixed concrete wall, and relationship to coupling station (TAN-620) and rail track. Ralph M. Parson 902-4-ANP-624-A 329. Date: February 1954. Approved by INEEL Classification Office for public release. INEEL Index code no. 035-0624-00-693-106911 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

Stem/Progenitor Cell–Mediated De Novo Regeneration of Dental Pulp with Newly Deposited Continuous Layer of Dentin in an In Vivo Model

PubMed Central

Yamaza, Takayoshi; Shea, Lonnie D.; Djouad, Farida; Kuhn, Nastaran Z.; Tuan, Rocky S.; Shi, Songtao

2010-01-01

The ultimate goal of this study is to regenerate lost dental pulp and dentin via stem/progenitor cell–based approaches and tissue engineering technologies. In this study, we tested the possibility of regenerating vascularized human dental pulp in emptied root canal space and producing new dentin on existing dentinal walls using a stem/progenitor cell–mediated approach with a human root fragment and an immunocompromised mouse model. Stem/progenitor cells from apical papilla and dental pulp stem cells were isolated, characterized, seeded onto synthetic scaffolds consisting of poly-D,L-lactide/glycolide, inserted into the tooth fragments, and transplanted into mice. Our results showed that the root canal space was filled entirely by a pulp-like tissue with well-established vascularity. In addition, a continuous layer of dentin-like tissue was deposited onto the canal dentinal wall. This dentin-like structure appeared to be produced by a layer of newly formed odontoblast-like cells expressing dentin sialophosphoprotein, bone sialoprotein, alkaline phosphatase, and CD105. The cells in regenerated pulp-like tissue reacted positively to anti-human mitochondria antibodies, indicating their human origin. This study provides the first evidence showing that pulp-like tissue can be regenerated de novo in emptied root canal space by stem cells from apical papilla and dental pulp stem cells that give rise to odontoblast-like cells producing dentin-like tissue on existing dentinal walls. PMID:19737072

Engineering nanoscale surface features to sustain microparticle rolling in flow.

PubMed

Kalasin, Surachate; Santore, Maria M

2015-05-26

Nanoscopic features of channel walls are often engineered to facilitate microfluidic transport, for instance when surface charge enables electro-osmosis or when grooves drive mixing. The dynamic or rolling adhesion of flowing microparticles on a channel wall holds potential to accomplish particle sorting or to selectively transfer reactive species or signals between the wall and flowing particles. Inspired by cell rolling under the direction of adhesion molecules called selectins, we present an engineered platform in which the rolling of flowing microparticles is sustained through the incorporation of entirely synthetic, discrete, nanoscale, attractive features into the nonadhesive (electrostatically repulsive) surface of a flow channel. Focusing on one example or type of nanoscale feature and probing the impact of broad systematic variations in surface feature loading and processing parameters, this study demonstrates how relatively flat, weakly adhesive nanoscale features, positioned with average spacings on the order of tens of nanometers, can produce sustained microparticle rolling. We further demonstrate how the rolling velocity and travel distance depend on flow and surface design. We identify classes of related surfaces that fail to support rolling and present a state space that identifies combinations of surface and processing variables corresponding to transitions between rolling, free particle motion, and arrest. Finally we identify combinations of parameters (surface length scales, particle size, flow rates) where particles can be manipulated with size-selectivity.

Identification of Cell Wall Synthesis Regulatory Genes Controlling Biomass Characteristics and Yield in Rice (Oryza Sativa)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Peng, Zhaohua PEng; Ronald, Palmela; Wang, Guo-Liang

This project aims to identify the regulatory genes of rice cell wall synthesis pathways using a cell wall removal and regeneration system. We completed the gene expression profiling studies following the time course from cell wall removal to cell wall regeneration in rice suspension cells. We also completed, total proteome, nuclear subproteome and histone modification studies following the course from cell wall removal and cell wall regeneration process. A large number of differentially expressed regulatory genes and proteins were identified. Meanwhile, we generated RNAi and over-expression transgenic rice for 45 genes with at least 10 independent transgenic lines for eachmore » gene. In addition, we ordered T-DNA and transposon insertion mutants for 60 genes from Korea, Japan, and France and characterized the mutants. Overall, we have mutants and transgenic lines for over 90 genes, exceeded our proposed goal of generating mutants for 50 genes. Interesting Discoveries a) Cell wall re-synthesis in protoplasts may involve a novel cell wall synthesis mechanism. The synthesis of the primary cell wall is initiated in late cytokinesis with further modification during cell expansion. Phragmoplast plays an essential role in cell wall synthesis. It services as a scaffold for building the cell plate and formation of a new cell wall. Only one phragmoplast and one new cell wall is produced for each dividing cell. When the cell wall was removed enzymatically, we found that cell wall re-synthesis started from multiple locations simultaneously, suggesting that a novel mechanism is involved in cell wall re-synthesis. This observation raised many interesting questions, such as how the starting sites of cell wall synthesis are determined, whether phragmoplast and cell plate like structures are involved in cell wall re-synthesis, and more importantly whether the same set of enzymes and apparatus are used in cell wall re-synthesis as during cytokinesis. Given that many known cell wall synthesis pathway genes are induced by removal of cell wall, some cell wall synthesis apparatus must be shared in both cases. The cell wall re-synthesis mechanism may have broad application because our preliminary assay indicates that the cell wall characteristics are highly different from those produced during cytokinesis. A thorough understanding on the regulation of cell wall re-synthesis may lead to improvement of cell wall characteristics. b) Removal of cell wall results in chromatin decondensation Another interesting observation was that removal of cell wall was associated with substantial chromatin change. Our DNA DAPI stain, chromatin MNase digestion, histone modification proteomics, protein differential expression analysis, and DNA oligo array studies all supported that substantial chromatin change was associated with removal of cell wall treatment. It is still under investigation if the chromatin change is associated with activation of cell wall synthesis genes, in which chromatin remodeling is required. Another possibility is that the cell wall is required for stabilizing the chromatin structure in plant cells. Given that spindle fiber is directly connected with both chromatin structure and cell wall synthesis, it is possible that there is an intrinsic connection between cell wall and chromatin.« less

Fuel injector for use in a gas turbine engine

DOEpatents

Wiebe, David J.

2012-10-09

A fuel injector in a combustor apparatus of a gas turbine engine. An outer wall of the injector defines an interior volume in which an intermediate wall is disposed. A first gap is formed between the outer wall and the intermediate wall. The intermediate wall defines an internal volume in which an inner wall is disposed. A second gap is formed between the intermediate wall and the inner wall. The second gap receives cooling fluid that cools the injector. The cooling fluid provides convective cooling to the intermediate wall as it flows within the second gap. The cooling fluid also flows through apertures in the intermediate wall into the first gap where it provides impingement cooling to the outer wall and provides convective cooling to the outer wall. The inner wall defines a passageway that delivers fuel into a liner downstream from a main combustion zone.

Architecture of dermatophyte cell Walls: Electron microscopic and biochemical analysis

NASA Technical Reports Server (NTRS)

Nozawa, Y.; Kitajima, Y.

1984-01-01

A review with 83 references on the cell wall structure of dermatophytes is presented. Topics discussed include separation and preparation of cell walls; microstructure of cell walls by electron microscopy; chemical composition of cell walls; structural model of cell walls; and morphological structure of cell walls.

Arabidopsis Regenerating Protoplast: A Powerful Model System for Combining the Proteomics of Cell Wall Proteins and the Visualization of Cell Wall Dynamics

PubMed Central

Yokoyama, Ryusuke; Kuki, Hiroaki; Kuroha, Takeshi; Nishitani, Kazuhiko

2016-01-01

The development of a range of sub-proteomic approaches to the plant cell wall has identified many of the cell wall proteins. However, it remains difficult to elucidate the precise biological role of each protein and the cell wall dynamics driven by their actions. The plant protoplast provides an excellent means not only for characterizing cell wall proteins, but also for visualizing the dynamics of cell wall regeneration, during which cell wall proteins are secreted. It therefore offers a unique opportunity to investigate the de novo construction process of the cell wall. This review deals with sub-proteomic approaches to the plant cell wall through the use of protoplasts, a methodology that will provide the basis for further exploration of cell wall proteins and cell wall dynamics. PMID:28248244

Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response

DOE PAGES

Tsai, Alex Yi-Lin; Chan, Kin; Ho, Chi-Yip; ...

2017-03-02

The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to themore » hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that transgenic expression of microbial cell wall modifying enzymes induces transcriptome responses similar to those observed in defense responses.« less

Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsai, Alex Yi-Lin; Chan, Kin; Ho, Chi-Yip

The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to themore » hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that transgenic expression of microbial cell wall modifying enzymes induces transcriptome responses similar to those observed in defense responses.« less

Characterization of the Sclerotinia sclerotiorum cell wall proteome.

PubMed

Liu, Longzhou; Free, Stephen J

2016-08-01

We used a proteomic analysis to identify cell wall proteins released from Sclerotinia sclerotiorum hyphal and sclerotial cell walls via a trifluoromethanesulfonic acid (TFMS) digestion. Cell walls from hyphae grown in Vogel's glucose medium (a synthetic medium lacking plant materials), from hyphae grown in potato dextrose broth and from sclerotia produced on potato dextrose agar were used in the analysis. Under the conditions used, TFMS digests the glycosidic linkages in the cell walls to release intact cell wall proteins. The analysis identified 24 glycosylphosphatidylinositol (GPI)-anchored cell wall proteins and 30 non-GPI-anchored cell wall proteins. We found that the cell walls contained an array of cell wall biosynthetic enzymes similar to those found in the cell walls of other fungi. When comparing the proteins in hyphal cell walls grown in potato dextrose broth with those in hyphal cell walls grown in the absence of plant material, it was found that a core group of cell wall biosynthetic proteins and some proteins associated with pathogenicity (secreted cellulases, pectin lyases, glucosidases and proteases) were expressed in both types of hyphae. The hyphae grown in potato dextrose broth contained a number of additional proteins (laccases, oxalate decarboxylase, peroxidase, polysaccharide deacetylase and several proteins unique to Sclerotinia and Botrytis) that might facilitate growth on a plant host. A comparison of the proteins in the sclerotial cell wall with the proteins in the hyphal cell wall demonstrated that sclerotia formation is not marked by a major shift in the composition of cell wall protein. We found that the S. sclerotiorum cell walls contained 11 cell wall proteins that were encoded only in Sclerotinia and Botrytis genomes. © 2015 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.

The valine and lysine residues in the conserved FxVTxK motif are important for the function of phylogenetically distant plant cellulose synthases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Slabaugh, Erin; Scavuzzo-Duggan, Tess; Chaves, Arielle

2015-12-08

Cellulose synthases (CESAs) synthesize the β-1,4-glucan chains that coalesce to form cellulose microfibrils in plant cell walls. In addition to a large cytosolic (catalytic) domain, CESAs have eight predicted transmembrane helices (TMHs). However, analogous to the structure of BcsA, a bacterial CESA, predicted TMH5 in CESA may instead be an interfacial helix. This would place the conserved FxVTxK motif in the plant cell cytosol where it could function as a substrate-gating loop as occurs in BcsA. To define the functional importance of the CESA region containing FxVTxK, we tested five parallel mutations in Arabidopsis thaliana CESA1 and Physcomitrella patens CESA5more » in complementation assays of the relevant cesa mutants. In both organisms, the substitution of the valine or lysine residues in FxVTxK severely affected CESA function. In Arabidopsis roots, both changes were correlated with lower cellulose anisotropy, as revealed by Pontamine Fast Scarlet. Analysis of hypocotyl inner cell wall layers by atomic force microscopy showed that two altered versions of Atcesa1 could rescue cell wall phenotypes observed in the mutant background line. Overall, the data show that the FxVTxK motif is functionally important in two phylogenetically distant plant CESAs. The results show that Physcomitrella provides an efficient model for assessing the effects of engineered CESA mutations affecting primary cell wall synthesis and that diverse testing systems can lead to nuanced insights into CESA structure–function relationships. Although CESA membrane topology needs to be experimentally determined, the results support the possibility that the FxVTxK region functions similarly in CESA and BcsA.« less

Cellulose-hemicellulose interaction in wood secondary cell-wall

NASA Astrophysics Data System (ADS)

Zhang, Ning; Li, Shi; Xiong, Liming; Hong, Yu; Chen, Youping

2015-12-01

The wood cell wall features a tough and relatively rigid fiber reinforced composite structure. It acts as a pressure vessel, offering protection against mechanical stress. Cellulose microfibrils, hemicellulose and amorphous lignin are the three major components of wood. The structure of secondary cell wall could be imagined as the same as reinforced concrete, in which cellulose microfibrils acts as reinforcing steel bar and hemicellulose-lignin matrices act as the concrete. Therefore, the interface between cellulose and hemicellulose/lignin plays a significant role in determine the mechanical behavior of wood secondary cell wall. To this end, we present a molecular dynamics (MD) simulation study attempting to quantify the strength of the interface between cellulose microfibrils and hemicellulose. Since hemicellulose binds with adjacent cellulose microfibrils in various patterns, the atomistic models of hemicellulose-cellulose composites with three typical binding modes, i.e. bridge, loop and random binding modes are constructed. The effect of the shape of hemicellulose chain on the strength of hemicellulose-cellulose composites under shear loadings is investigated. The contact area as well as hydrogen bonds between cellulose and hemicellulose, together with the covalent bonds in backbone of hemicellulose chain are found to be the controlling parameters which determine the strength of the interfaces in the composite system. For the bridge binding model, the effect of shear loading direction on the strength of the cellulose material is also studied. The obtained results suggest that the shear strength of wood-inspired engineering composites can be optimized through maximizing the formations of the contributing hydrogen bonds between cellulose and hemicellulose.

Critical review on the mechanisms of maturation stress generation in trees.

PubMed

Alméras, Tancrède; Clair, Bruno

2016-09-01

Trees control their posture by generating asymmetric mechanical stress around the periphery of the trunk or branches. This stress is produced in wood during the maturation of the cell wall. When the need for reaction is high, it is accompanied by strong changes in cell organization and composition called reaction wood, namely compression wood in gymnosperms and tension wood in angiosperms. The process by which stress is generated in the cell wall during its formation is not yet known, and various hypothetical mechanisms have been proposed in the literature. Here we aim at discriminating between these models. First, we summarize current knowledge about reaction wood structure, state and behaviour relevant to the understanding of maturation stress generation. Then, the mechanisms proposed in the literature are listed and discussed in order to identify which can be rejected based on their inconsistency with current knowledge at the frontier between plant science and mechanical engineering. © 2016 The Author(s).

An assessment of CFD-based wall heat transfer models in piston engines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sircar, Arpan; Paul, Chandan; Ferreyro-Fernandez, Sebastian

The lack of accurate submodels for in-cylinder heat transfer has been identified as a key shortcoming in developing truly predictive, physics-based computational fluid dynamics (CFD) models that can be used to develop combustion systems for advanced high-efficiency, low-emissions engines. Only recently have experimental methods become available that enable accurate near-wall measurements to enhance simulation capability via advancing models. Initial results show crank-angle dependent discrepancies with respect to previously used boundary-layer models of up to 100%. However, available experimental data is quite sparse (only few data points on engine walls) and limited (available measurements are those of heat flux only). Predictivemore » submodels are needed for medium-resolution ("engineering") LES and for unsteady Reynolds-averaged simulations (URANS). Recently, some research groups have performed DNS studies on engine-relevant conditions using simple geometries. These provide very useful data for benchmarking wall heat transfer models under such conditions. Further, a number of new and more sophisticated models have also become available in the literature which account for these engine-like conditions. Some of these have been incorporated while others of a more complex nature, which include solving additional partial differential equations (PDEs) within the thin boundary layer near the wall, are underway. These models will then be tested against the available DNS/experimental data in both SI (spark-ignition) and CI (compression-ignition) engines.« less

Nuclear Rocket Test Facility Decommissioning Including Controlled Explosive Demolition of a Neutron-Activated Shield Wall

DOE Office of Scientific and Technical Information (OSTI.GOV)

Michael Kruzic

2007-09-01

Located in Area 25 of the Nevada Test Site, the Test Cell A Facility was used in the 1960s for the testing of nuclear rocket engines, as part of the Nuclear Rocket Development Program. The facility was decontaminated and decommissioned (D&D) in 2005 using the Streamlined Approach For Environmental Restoration (SAFER) process, under the Federal Facilities Agreement and Consent Order (FFACO). Utilities and process piping were verified void of contents, hazardous materials were removed, concrete with removable contamination decontaminated, large sections mechanically demolished, and the remaining five-foot, five-inch thick radiologically-activated reinforced concrete shield wall demolished using open-air controlled explosive demolitionmore » (CED). CED of the shield wall was closely monitored and resulted in no radiological exposure or atmospheric release.« less

ETR BUILDING, TRA642, INTERIOR. BASEMENT. CORRIDOR ALONG WEST WALL OF ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

ETR BUILDING, TRA-642, INTERIOR. BASEMENT. CORRIDOR ALONG WEST WALL OF BUILDING, WHICH IS AT RIGHT OF VIEW. AUDIO ALARM IS ALONG WALL AT RIGHT. CAMERA FACES SOUTH. INL NEGATIVE NO. HD46-30-1. Mike Crane, Photographer, 2/2005 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

The Role of Auxin in Cell Wall Expansion

PubMed Central

2018-01-01

Plant cells are surrounded by cell walls, which are dynamic structures displaying a strictly regulated balance between rigidity and flexibility. Walls are fairly rigid to provide support and protection, but also extensible, to allow cell growth, which is triggered by a high intracellular turgor pressure. Wall properties regulate the differential growth of the cell, resulting in a diversity of cell sizes and shapes. The plant hormone auxin is well known to stimulate cell elongation via increasing wall extensibility. Auxin participates in the regulation of cell wall properties by inducing wall loosening. Here, we review what is known on cell wall property regulation by auxin. We focus particularly on the auxin role during cell expansion linked directly to cell wall modifications. We also analyze downstream targets of transcriptional auxin signaling, which are related to the cell wall and could be linked to acid growth and the action of wall-loosening proteins. All together, this update elucidates the connection between hormonal signaling and cell wall synthesis and deposition. PMID:29565829

The Role of Auxin in Cell Wall Expansion.

PubMed

Majda, Mateusz; Robert, Stéphanie

2018-03-22

Plant cells are surrounded by cell walls, which are dynamic structures displaying a strictly regulated balance between rigidity and flexibility. Walls are fairly rigid to provide support and protection, but also extensible, to allow cell growth, which is triggered by a high intracellular turgor pressure. Wall properties regulate the differential growth of the cell, resulting in a diversity of cell sizes and shapes. The plant hormone auxin is well known to stimulate cell elongation via increasing wall extensibility. Auxin participates in the regulation of cell wall properties by inducing wall loosening. Here, we review what is known on cell wall property regulation by auxin. We focus particularly on the auxin role during cell expansion linked directly to cell wall modifications. We also analyze downstream targets of transcriptional auxin signaling, which are related to the cell wall and could be linked to acid growth and the action of wall-loosening proteins. All together, this update elucidates the connection between hormonal signaling and cell wall synthesis and deposition.

Correlation between vortices and wall shear stress in a curved artery model under pulsatile flow conditions

NASA Astrophysics Data System (ADS)

Cox, Christopher; Plesniak, Michael W.

2017-11-01

One of the most physiologically relevant factors within the cardiovascular system is the wall shear stress. The wall shear stress affects endothelial cells via mechanotransduction and atherosclerotic regions are strongly correlated with curvature and branching in the human vasculature, where the shear stress is both oscillatory and multidirectional. Also, the combined effect of curvature and pulsatility in cardiovascular flows produces unsteady vortices. In this work, our goal is to assess the correlation between multiple vortex pairs and wall shear stress. To accomplish this, we use an in-house high-order flux reconstruction Navier-Stokes solver to simulate pulsatile flow of a Newtonian blood-analog fluid through a rigid 180° curved artery model. We use a physiologically relevant flow rate and generate results using both fully developed and uniform entrance conditions, the latter motivated by the fact that flow upstream to a curved artery may not be fully developed. Under these two inflow conditions, we characterize the evolution of various vortex pairs and their subsequent effect on several wall shear stress metrics. Supported by GW Center for Biomimetics and Bioinspired Engineering.

Empowering Marine Corps System Administrators: Taxonomy of Training

DTIC Science & Technology

2004-03-01

of Systems and Engineering Management Graduate School of Engineering and Management Air Force Institute of Technology Air University Air...Information Assurance ................................................................................ 16 2.8 Attacks, Social Engineering & Online Users... drawbridge , outer castle walls, inner castle walls and the keep. No single form of defense is foolproof, thus each layer compensates for deficiencies

Transpiration Cooling Experiment

NASA Technical Reports Server (NTRS)

Song, Kyo D.; Ries, Heidi R.; Scotti, Stephen J.; Choi, Sang H.

1997-01-01

The transpiration cooling method was considered for a scram-jet engine to accommodate thermally the situation where a very high heat flux (200 Btu/sq. ft sec) from hydrogen fuel combustion process is imposed to the engine walls. In a scram-jet engine, a small portion of hydrogen fuel passes through the porous walls of the engine combustor to cool the engine walls and at the same time the rest passes along combustion chamber walls and is preheated. Such a regenerative system promises simultaneously cooling of engine combustor and preheating the cryogenic fuel. In the experiment, an optical heating method was used to provide a heat flux of 200 Btu/sq. ft sec to the cylindrical surface of a porous stainless steel specimen which carried helium gas. The cooling efficiencies by transpiration were studied for specimens with various porosity. The experiments of various test specimens under high heat flux have revealed a phenomenon that chokes the medium flow when passing through a porous structure. This research includes the analysis of the system and a scaling conversion study that interprets the results from helium into the case when hydrogen medium is used.

Engineering a monolignol 4- O-methyltransferase with high selectivity for the condensed lignin precursor coniferyl alchohol

DOE PAGES

Cai, Yuanheng; Shanklin, John; Mohammad -Wadud Bhuiya; ...

2015-09-16

Lignin, a rigid biopolymer in plant cell walls, is derived from the oxidative polymerization of three monolignols. The composition of monolignol monomers dictates the degree of lignin condensation, reactivity, and thus the degradability of plant cell walls. Guaiacyl lignin is regarded as the condensed structural unit. Polymerization of lignin is initiated through the deprotonation of the para-hydroxyl group of monolignols. Therefore, preferentially modifying the para-hydroxyl of a specific monolignol to deprive its dehydrogenation propensity would disturb the formation of particular lignin subunits. Here, we test the hypothesis that specific remodeling the active site of a monolignol 4-O-methyltransferase would create anmore » enzyme that specifically methylates the condensed guaiacyl lignin precursor coniferyl alcohol. Combining crystal structural information with combinatorial active site saturation mutagenesis and starting with the engineered promiscuous enzyme, MOMT5 (T133L/E165I/F175I/F166W/H169F), we incrementally remodeled its substrate binding pocket by the addition of four substitutions, i.e. M26H, S30R, V33S, and T319M, yielding a mutant enzyme capable of discriminately etherifying the para-hydroxyl of coniferyl alcohol even in the presence of excess sinapyl alcohol. The engineered enzyme variant has a substantially reduced substrate binding pocket that imposes a clear steric hindrance thereby excluding bulkier lignin precursors. Lastly, the resulting enzyme variant represents an excellent candidate for modulating lignin composition and/or structure in planta.« less

Engineering a Monolignol 4-O-Methyltransferase with High Selectivity for the Condensed Lignin Precursor Coniferyl Alcohol*

PubMed Central

Cai, Yuanheng; Bhuiya, Mohammad-Wadud; Shanklin, John; Liu, Chang-Jun

2015-01-01

Lignin, a rigid biopolymer in plant cell walls, is derived from the oxidative polymerization of three monolignols. The composition of monolignol monomers dictates the degree of lignin condensation, reactivity, and thus the degradability of plant cell walls. Guaiacyl lignin is regarded as the condensed structural unit. Polymerization of lignin is initiated through the deprotonation of the para-hydroxyl group of monolignols. Therefore, preferentially modifying the para-hydroxyl of a specific monolignol to deprive its dehydrogenation propensity would disturb the formation of particular lignin subunits. Here, we test the hypothesis that specific remodeling the active site of a monolignol 4-O-methyltransferase would create an enzyme that specifically methylates the condensed guaiacyl lignin precursor coniferyl alcohol. Combining crystal structural information with combinatorial active site saturation mutagenesis and starting with the engineered promiscuous enzyme, MOMT5 (T133L/E165I/F175I/F166W/H169F), we incrementally remodeled its substrate binding pocket by the addition of four substitutions, i.e. M26H, S30R, V33S, and T319M, yielding a mutant enzyme capable of discriminately etherifying the para-hydroxyl of coniferyl alcohol even in the presence of excess sinapyl alcohol. The engineered enzyme variant has a substantially reduced substrate binding pocket that imposes a clear steric hindrance thereby excluding bulkier lignin precursors. The resulting enzyme variant represents an excellent candidate for modulating lignin composition and/or structure in planta. PMID:26378240

Droplet-Wall/Film Impact in IC Engine Applications

DTIC Science & Technology

2017-08-14

Report: Droplet-Wall/Film Impact in IC Engine Applications (ARO Topic 1.4.1 under ARO’s Dr. Ralph A. Anthenien) The views, opinions and/or findings...in IC Engine Applications (ARO Topic 1.4.1 under ARO’s Dr. Ralph A. Anthenien) Report Term: 0-Other Email: cklaw@princeton.edu Distribution Statement...associated with spraying in internal combustion engines (ICEs). Fuels sprayed inside engines can impact with the internal surfaces and thus not only

Bioreactors Drive Advances in Tissue Engineering

NASA Technical Reports Server (NTRS)

2012-01-01

It was an unlikely moment for inspiration. Engineers David Wolf and Ray Schwarz stopped by their lab around midday. Wolf, of Johnson Space Center, and Schwarz, with NASA contractor Krug Life Sciences (now Wyle Laboratories Inc.), were part of a team tasked with developing a unique technology with the potential to enhance medical research. But that wasn t the focus at the moment: The pair was rounding up colleagues interested in grabbing some lunch. One of the lab s other Krug engineers, Tinh Trinh, was doing something that made Wolf forget about food. Trinh was toying with an electric drill. He had stuck the barrel of a syringe on the bit; it spun with a high-pitched whirr when he squeezed the drill s trigger. At the time, a multidisciplinary team of engineers and biologists including Wolf, Schwarz, Trinh, and project manager Charles D. Anderson, who formerly led the recovery of the Apollo capsules after splashdown and now worked for Krug was pursuing the development of a technology called a bioreactor, a cylindrical device used to culture human cells. The team s immediate goal was to grow human kidney cells to produce erythropoietin, a hormone that regulates red blood cell production and can be used to treat anemia. But there was a major barrier to the technology s success: Moving the liquid growth media to keep it from stagnating resulted in turbulent conditions that damaged the delicate cells, causing them to quickly die. The team was looking forward to testing the bioreactor in space, hoping the device would perform more effectively in microgravity. But on January 28, 1986, the Space Shuttle Challenger broke apart shortly after launch, killing its seven crewmembers. The subsequent grounding of the shuttle fleet had left researchers with no access to space, and thus no way to study the effects of microgravity on human cells. As Wolf looked from Trinh s syringe-capped drill to where the bioreactor sat on a workbench, he suddenly saw a possible solution to both problems. It dawned on me that rotating the wall of the reactor would solve one of our fundamental fluid mechanical problems, specifically by removing the velocity gradient of the tissue culture fluid media near the reactor s walls, says Wolf. It looked as though it would allow us to suspend the growing cells within the reactor without introducing turbulent fluid mechanical conditions.

Cell Wall Remodeling Enzymes Modulate Fungal Cell Wall Elasticity and Osmotic Stress Resistance

PubMed Central

Ene, Iuliana V.; Walker, Louise A.; Schiavone, Marion; Lee, Keunsook K.; Martin-Yken, Hélène; Dague, Etienne; Gow, Neil A. R.; Munro, Carol A.

2015-01-01

ABSTRACT The fungal cell wall confers cell morphology and protection against environmental insults. For fungal pathogens, the cell wall is a key immunological modulator and an ideal therapeutic target. Yeast cell walls possess an inner matrix of interlinked β-glucan and chitin that is thought to provide tensile strength and rigidity. Yeast cells remodel their walls over time in response to environmental change, a process controlled by evolutionarily conserved stress (Hog1) and cell integrity (Mkc1, Cek1) signaling pathways. These mitogen-activated protein kinase (MAPK) pathways modulate cell wall gene expression, leading to the construction of a new, modified cell wall. We show that the cell wall is not rigid but elastic, displaying rapid structural realignments that impact survival following osmotic shock. Lactate-grown Candida albicans cells are more resistant to hyperosmotic shock than glucose-grown cells. We show that this elevated resistance is not dependent on Hog1 or Mkc1 signaling and that most cell death occurs within 10 min of osmotic shock. Sudden decreases in cell volume drive rapid increases in cell wall thickness. The elevated stress resistance of lactate-grown cells correlates with reduced cell wall elasticity, reflected in slower changes in cell volume following hyperosmotic shock. The cell wall elasticity of lactate-grown cells is increased by a triple mutation that inactivates the Crh family of cell wall cross-linking enzymes, leading to increased sensitivity to hyperosmotic shock. Overexpressing Crh family members in glucose-grown cells reduces cell wall elasticity, providing partial protection against hyperosmotic shock. These changes correlate with structural realignment of the cell wall and with the ability of cells to withstand osmotic shock. PMID:26220968

Simulated microgravity does not alter epithelial cell adhesion to matrix and other molecules

NASA Technical Reports Server (NTRS)

Jessup, J. M.; Brown, K.; Ishii, S.; Ford, R.; Goodwin, T. J.; Spaulding, G.

1994-01-01

Microgravity has advantages for the cultivation of tissues with high fidelity; however, tissue formation requires cellular recognition and adhesion. We tested the hypothesis that simulated microgravity does not affect cell adhesion. Human colorectal carcinoma cells were cultured in the NASA Rotating Wall Vessel (RWV) under low shear stress with randomization of the gravity vector that simulates microgravity. After 6 - 7 days, cells were assayed for binding to various substrates and compared to cells grown in standard tissue culture flasks and static suspension cultures. The RWV cultures bound as well to basement membrane proteins and to Carcinoembryonic Antigen (CEA), an intercellular adhesion molecule, as control cultures did. Thus, microgravity does not alter epithelial cell adhesion and may be useful for tissue engineering.

Central Control Room in the Engine Research Building

NASA Image and Video Library

1968-11-21

Operators in the Engine Research Building’s Central Control Room at the National Aeronautics and Space Administration (NASA) Lewis Research Center. The massive 4.25-acre Engine Research Building contains dozens of test cells, test stands, and altitude chambers. A powerful a collection of compressors and exhausters located in the central portion of the basement provides process air and exhaust for these test areas. This system is connected to similar process air systems in the laboratory’s other large test facilities. The Central Control Room coordinates this activity and communicates with the local utilities. The panels on the wall contain schematics with indicator lights and instrumentation for the atmospheric exhaust, altitude exhaust, refrigerated air, and process air systems. The process air equipment included twelve exhausters, four compressors, refrigeration system, cooling water, and an exhaust system. The operators in the control room kept in contact with engineers running the process air system and those conducting the tests in the test cells. The operators also coordinated with the local power companies to make sure enough electricity was available to operate the powerful compressors and exhausters.

Cell Wall Remodeling Enzymes Modulate Fungal Cell Wall Elasticity and Osmotic Stress Resistance.

PubMed

Ene, Iuliana V; Walker, Louise A; Schiavone, Marion; Lee, Keunsook K; Martin-Yken, Hélène; Dague, Etienne; Gow, Neil A R; Munro, Carol A; Brown, Alistair J P

2015-07-28

The fungal cell wall confers cell morphology and protection against environmental insults. For fungal pathogens, the cell wall is a key immunological modulator and an ideal therapeutic target. Yeast cell walls possess an inner matrix of interlinked β-glucan and chitin that is thought to provide tensile strength and rigidity. Yeast cells remodel their walls over time in response to environmental change, a process controlled by evolutionarily conserved stress (Hog1) and cell integrity (Mkc1, Cek1) signaling pathways. These mitogen-activated protein kinase (MAPK) pathways modulate cell wall gene expression, leading to the construction of a new, modified cell wall. We show that the cell wall is not rigid but elastic, displaying rapid structural realignments that impact survival following osmotic shock. Lactate-grown Candida albicans cells are more resistant to hyperosmotic shock than glucose-grown cells. We show that this elevated resistance is not dependent on Hog1 or Mkc1 signaling and that most cell death occurs within 10 min of osmotic shock. Sudden decreases in cell volume drive rapid increases in cell wall thickness. The elevated stress resistance of lactate-grown cells correlates with reduced cell wall elasticity, reflected in slower changes in cell volume following hyperosmotic shock. The cell wall elasticity of lactate-grown cells is increased by a triple mutation that inactivates the Crh family of cell wall cross-linking enzymes, leading to increased sensitivity to hyperosmotic shock. Overexpressing Crh family members in glucose-grown cells reduces cell wall elasticity, providing partial protection against hyperosmotic shock. These changes correlate with structural realignment of the cell wall and with the ability of cells to withstand osmotic shock. The C. albicans cell wall is the first line of defense against external insults, the site of immune recognition by the host, and an attractive target for antifungal therapy. Its tensile strength is conferred by a network of cell wall polysaccharides, which are remodeled in response to growth conditions and environmental stress. However, little is known about how cell wall elasticity is regulated and how it affects adaptation to stresses such as sudden changes in osmolarity. We show that elasticity is critical for survival under conditions of osmotic shock, before stress signaling pathways have time to induce gene expression and drive glycerol accumulation. Critical cell wall remodeling enzymes control cell wall flexibility, and its regulation is strongly dependent on host nutritional inputs. We also demonstrate an entirely new level of cell wall dynamism, where significant architectural changes and structural realignment occur within seconds of an osmotic shock. Copyright © 2015 Ene et al.

Stomatal Opening: The Role of Cell-Wall Mechanical Anisotropy and Its Analytical Relations to the Bio-composite Characteristics

PubMed Central

Marom, Ziv; Shtein, Ilana; Bar-On, Benny

2017-01-01

Stomata are pores on the leaf surface, which are formed by a pair of curved, tubular guard cells; an increase in turgor pressure deforms the guard cells, resulting in the opening of the stomata. Recent studies employed numerical simulations, based on experimental data, to analyze the effects of various structural, chemical, and mechanical features of the guard cells on the stomatal opening characteristics; these studies all support the well-known qualitative observation that the mechanical anisotropy of the guard cells plays a critical role in stomatal opening. Here, we propose a computationally based analytical model that quantitatively establishes the relations between the degree of anisotropy of the guard cell, the bio-composite constituents of the cell wall, and the aperture and area of stomatal opening. The model introduces two non-dimensional key parameters that dominate the guard cell deformations—the inflation driving force and the anisotropy ratio—and it serves as a generic framework that is not limited to specific plant species. The modeling predictions are in line with a wide range of previous experimental studies, and its analytical formulation sheds new light on the relations between the structure, mechanics, and function of stomata. Moreover, the model provides an analytical tool to back-calculate the elastic characteristics of the matrix that composes the guard cell walls, which, to the best of our knowledge, cannot be probed by direct nano-mechanical experiments; indeed, the estimations of our model are in good agreement with recently published results of independent numerical optimization schemes. The emerging insights from the stomatal structure-mechanics “design guidelines” may promote the development of miniature, yet complex, multiscale composite actuation mechanisms for future engineering platforms. PMID:29312365

How do plants enlarge? A balancing act; Final report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boyer, J.S.

1996-12-31

Cells of plants are surrounded by strong walls that prevent rupture from internal pressures that can be two or three times that of an automobile tire. In this way, the walls protect the cytoplasm. However, at the same time, the cells can enlarge as they grow. How this balancing act works and how it enlarges the plant were the subject of a recent conference at the University of Delaware in Lewes. The aim was to identify areas for future research that could explain the enlargement of whole plants. There is a large practical need to predict and modify plant enlargementmore » but the additional processes that overlie the molecular ones need to be integrated with the molecular information before a picture will emerge. How best to accomplish this involved input from cross-disciplinary areas in biomechanics, physics and engineering as well as molecular biology, biochemistry and ultrastructure.« less

Designer biomass for next-generation biorefineries: leveraging recent insights into xylan structure and biosynthesis.

PubMed

Smith, Peter J; Wang, Hsin-Tzu; York, William S; Peña, Maria J; Urbanowicz, Breeanna R

2017-01-01

Xylans are the most abundant noncellulosic polysaccharides in lignified secondary cell walls of woody dicots and in both primary and secondary cell walls of grasses. These polysaccharides, which comprise 20-35% of terrestrial biomass, present major challenges for the efficient microbial bioconversion of lignocellulosic feedstocks to fuels and other value-added products. Xylans play a significant role in the recalcitrance of biomass to degradation, and their bioconversion requires metabolic pathways that are distinct from those used to metabolize cellulose. In this review, we discuss the key differences in the structural features of xylans across diverse plant species, how these features affect their interactions with cellulose and lignin, and recent developments in understanding their biosynthesis. In particular, we focus on how the combined structural and biosynthetic knowledge can be used as a basis for biomass engineering aimed at developing crops that are better suited as feedstocks for the bioconversion industry.

Energy efficient engine high-pressure turbine supersonic cascade technology report

NASA Technical Reports Server (NTRS)

Kopper, F. C.; Milano, R.; Davis, R. L.; Dring, R. P.; Stoeffler, R. C.

1981-01-01

The performance of two vane endwall geometries and three blade sections for the high-pressure turbine was evaluated in terms of the efficiency requirements of the Energy Efficient Engine high-pressure turbine component. The van endwall designs featured a straight wall and S-wall configuration. The blade designs included a base blade, straightback blade, and overcambered blade. Test results indicated that the S-wall vane configuration and the base blade configuration offered the most promising performance characteristics for the Energy Efficient Engine high-pressure turbine component.

Cooled airfoil in a turbine engine

DOEpatents

Vitt, Paul H; Kemp, David A; Lee, Ching-Pang; Marra, John J

2015-04-21

An airfoil in a gas turbine engine includes an outer wall and an inner wall. The outer wall includes a leading edge, a trailing edge opposed from the leading edge in a chordal direction, a pressure side, and a suction side. The inner wall is coupled to the outer wall at a single chordal location and includes portions spaced from the pressure and suction sides of the outer wall so as to form first and second gaps between the inner wall and the respective pressure and suction sides. The inner wall defines a chamber therein and includes openings that provide fluid communication between the respective gaps and the chamber. The gaps receive cooling fluid that provides cooling to the outer wall as it flows through the gaps. The cooling fluid, after traversing at least substantial portions of the gaps, passes into the chamber through the openings in the inner wall.

3D Printed Vascular Networks Enhance Viability in High-Volume Perfusion Bioreactor.

PubMed

Ball, Owen; Nguyen, Bao-Ngoc B; Placone, Jesse K; Fisher, John P

2016-12-01

There is a significant clinical need for engineered bone graft substitutes that can quickly, effectively, and safely repair large segmental bone defects. One emerging field of interest involves the growth of engineered bone tissue in vitro within bioreactors, the most promising of which are perfusion bioreactors. Using bioreactor systems, tissue engineered bone constructs can be fabricated in vitro. However, these engineered constructs lack inherent vasculature and once implanted, quickly develop a necrotic core, where no nutrient exchange occurs. Here, we utilized COMSOL modeling to predict oxygen diffusion gradients throughout aggregated alginate constructs, which allowed for the computer-aided design of printable vascular networks, compatible with any large tissue engineered construct cultured in a perfusion bioreactor. We investigated the effect of 3D printed macroscale vascular networks with various porosities on the viability of human mesenchymal stem cells in vitro, using both gas-permeable, and non-gas permeable bioreactor growth chamber walls. Through the use of 3D printed vascular structures in conjunction with a tubular perfusion system bioreactor, cell viability was found to increase by as much as 50% in the core of these constructs, with in silico modeling predicting construct viability at steady state.

3D Printed Vascular Networks Enhance Viability in High-Volume Perfusion Bioreactor

PubMed Central

Ball, Owen; Nguyen, Bao-Ngoc B.; Placone, Jesse K.; Fisher, John P.

2016-01-01

There is a significant clinical need for engineered bone graft substitutes that can quickly, effectively, and safely repair large segmental bone defects. One emerging field of interest involves the growth of engineered bone tissue in vitro within bioreactors, the most promising of which are perfusion bioreactors. Using bioreactor systems, tissue engineered bone constructs can be fabricated in vitro. However, these engineered constructs lack inherent vasculature and once implanted, quickly develop a necrotic core, where no nutrient exchange occurs. Here, we utilized COMSOL modeling to predict oxygen diffusion gradients throughout aggregated alginate constructs, which allowed for the computer-aided design of printable vascular networks, compatible with any large tissue engineered construct cultured in a perfusion bioreactor. We investigated the effect of 3D printed macroscale vascular networks with various porosities on the viability of human mesenchymal stem cells in vitro, using both gas-permeable, and non-gas permeable bioreactor growth chamber walls. Through the use of 3D printed vascular structures in conjunction with a tubular perfusion system bioreactor, cell viability was found to increase by as much as 50% in the core of these constructs, with in silico modeling predicting construct viability at steady state. PMID:27272210

Cell wall targeted in planta iron accumulation enhances biomass conversion and seed iron concentration in Arabidopsis and rice

DOE PAGES

Yang, Haibing; Wei, Hui; Ma, Guojie; ...

2016-04-07

Conversion of nongrain biomass into liquid fuel is a sustainable approach to energy demands as global population increases. Previously, we showed that iron can act as a catalyst to enhance the degradation of lignocellulosic biomass for biofuel production. However, direct addition of iron catalysts to biomass pretreatment is diffusion-limited, would increase the cost and complexity of biorefinery unit operations and may have deleterious environmental impacts. Here, we show a new strategy for in planta accumulation of iron throughout the volume of the cell wall where iron acts as a catalyst in the deconstruction of lignocellulosic biomass. We engineered CBM-IBP fusionmore » polypeptides composed of a carbohydrate-binding module family 11 (CBM11) and an iron-binding peptide (IBP) for secretion into Arabidopsis and rice cell walls. CBM-IBP transformed Arabidopsis and rice plants show significant increases in iron accumulation and biomass conversion compared to respective controls. Further, CBM-IBP rice shows a 35% increase in seed iron concentration and a 40% increase in seed yield in greenhouse experiments. In conclusion, CBM-IBP rice potentially could be used to address iron deficiency, the most common and widespread nutritional disorder according to the World Health Organization.« less

Cell wall targeted in planta iron accumulation enhances biomass conversion and seed iron concentration in Arabidopsis and rice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Haibing; Wei, Hui; Ma, Guojie

Conversion of nongrain biomass into liquid fuel is a sustainable approach to energy demands as global population increases. Previously, we showed that iron can act as a catalyst to enhance the degradation of lignocellulosic biomass for biofuel production. However, direct addition of iron catalysts to biomass pretreatment is diffusion-limited, would increase the cost and complexity of biorefinery unit operations and may have deleterious environmental impacts. Here, we show a new strategy for in planta accumulation of iron throughout the volume of the cell wall where iron acts as a catalyst in the deconstruction of lignocellulosic biomass. We engineered CBM-IBP fusionmore » polypeptides composed of a carbohydrate-binding module family 11 (CBM11) and an iron-binding peptide (IBP) for secretion into Arabidopsis and rice cell walls. CBM-IBP transformed Arabidopsis and rice plants show significant increases in iron accumulation and biomass conversion compared to respective controls. Further, CBM-IBP rice shows a 35% increase in seed iron concentration and a 40% increase in seed yield in greenhouse experiments. CBM-IBP rice potentially could be used to address iron deficiency, the most common and widespread nutritional disorder according to the World Health Organization.« less

Cell wall targeted in planta iron accumulation enhances biomass conversion and seed iron concentration in Arabidopsis and rice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Haibing; Wei, Hui; Ma, Guojie

Conversion of nongrain biomass into liquid fuel is a sustainable approach to energy demands as global population increases. Previously, we showed that iron can act as a catalyst to enhance the degradation of lignocellulosic biomass for biofuel production. However, direct addition of iron catalysts to biomass pretreatment is diffusion-limited, would increase the cost and complexity of biorefinery unit operations and may have deleterious environmental impacts. Here, we show a new strategy for in planta accumulation of iron throughout the volume of the cell wall where iron acts as a catalyst in the deconstruction of lignocellulosic biomass. We engineered CBM-IBP fusionmore » polypeptides composed of a carbohydrate-binding module family 11 (CBM11) and an iron-binding peptide (IBP) for secretion into Arabidopsis and rice cell walls. CBM-IBP transformed Arabidopsis and rice plants show significant increases in iron accumulation and biomass conversion compared to respective controls. Further, CBM-IBP rice shows a 35% increase in seed iron concentration and a 40% increase in seed yield in greenhouse experiments. In conclusion, CBM-IBP rice potentially could be used to address iron deficiency, the most common and widespread nutritional disorder according to the World Health Organization.« less

Pullulan-based composite scaffolds for bone tissue engineering: Improved osteoconductivity by pore wall mineralization.

PubMed

Amrita; Arora, Aditya; Sharma, Poonam; Katti, Dhirendra S

2015-06-05

Porous hydrogels have been explored for bone tissue engineering; however their poor mechanical properties make them less suitable as bone graft substitutes. Since incorporation of fillers is a well-accepted method for improving mechanical properties of hydrogels, in this work pullulan hydrogels were reinforced with nano-crystalline hydroxyapatite (nHAp) (5 wt% nHAp in hydrogel) and poly(3-hydroxybutyrate) (PHB) fibers (3 wt% fibers in hydrogel) containing nHAp (3 wt% nHAp in fibers). Addition of these fillers to pullulan hydrogel improved compressive modulus of the scaffold by 10 fold. However, the hydrophilicity of pullulan did not support adhesion and spreading of cells. To overcome this limitation, porous composite scaffolds were modified using a double diffusion method that enabled deposition of hydroxyapatite on pore walls. This method resulted in rapid and uniform coating of HAp throughout the three-dimensional scaffolds which not only rendered them osteoconductive in vitro but also led to an improvement in their compressive modulus. These results demonstrate the potential of mineralized pullulan-based composite scaffolds in non-load bearing bone tissue engineering. Copyright © 2015 Elsevier Ltd. All rights reserved.

Differential histone modification and protein expression associated with cell wall removal and regeneration in rice (Oryza sativa).

PubMed

Tan, Feng; Zhang, Kangling; Mujahid, Hana; Verma, Desh Pal S; Peng, Zhaohua

2011-02-04

The cell wall is a critical extracellular structure that provides protection and structural support in plant cells. To study the biological function of the cell wall and the regulation of cell wall resynthesis, we examined cellular responses to enzymatic removal of the cell wall in rice (Oryza sativa) suspension cells using proteomic approaches. We find that removal of cell wall stimulates cell wall synthesis from multiple sites in protoplasts instead of from a single site as in cytokinesis. Nucleus DAPI stain and MNase digestion further show that removal of the cell wall is concomitant with substantial chromatin reorganization. Histone post-translational modification studies using both Western blots and isotope labeling assisted quantitative mass spectrometry analyses reveal that substantial histone modification changes, particularly H3K18(AC) and H3K23(AC), are associated with the removal and regeneration of the cell wall. Label-free quantitative proteome analyses further reveal that chromatin associated proteins undergo dramatic changes upon removal of the cell wall, along with cytoskeleton, cell wall metabolism, and stress-response proteins. This study demonstrates that cell wall removal is associated with substantial chromatin change and may lead to stimulation of cell wall synthesis using a novel mechanism.

WORKER STACKS GRAPHITE BLOCKS AGAINST INNER SOUTH WALL OF REACTOR. ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

WORKER STACKS GRAPHITE BLOCKS AGAINST INNER SOUTH WALL OF REACTOR. INL NEGATIVE NO. 3925. Unknown Photographer, 12/14/1951 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

On a turbulent wall model to predict hemolysis numerically in medical devices

NASA Astrophysics Data System (ADS)

Lee, Seunghun; Chang, Minwook; Kang, Seongwon; Hur, Nahmkeon; Kim, Wonjung

2017-11-01

Analyzing degradation of red blood cells is very important for medical devices with blood flows. The blood shear stress has been recognized as the most dominant factor for hemolysis in medical devices. Compared to laminar flows, turbulent flows have higher shear stress values in the regions near the wall. In case of predicting hemolysis numerically, this phenomenon can require a very fine mesh and large computational resources. In order to resolve this issue, the purpose of this study is to develop a turbulent wall model to predict the hemolysis more efficiently. In order to decrease the numerical error of hemolysis prediction in a coarse grid resolution, we divided the computational domain into two regions and applied different approaches to each region. In the near-wall region with a steep velocity gradient, an analytic approach using modeled velocity profile is applied to reduce a numerical error to allow a coarse grid resolution. We adopt the Van Driest law as a model for the mean velocity profile. In a region far from the wall, a regular numerical discretization is applied. The proposed turbulent wall model is evaluated for a few turbulent flows inside a cannula and centrifugal pumps. The results present that the proposed turbulent wall model for hemolysis improves the computational efficiency significantly for engineering applications. Corresponding author.

Enhancement of matrix production and cell proliferation in human annulus cells under bioreactor culture.

PubMed

Yang, Xinlin; Wang, Daidong; Hao, Jianrong; Gong, Meiqing; Arlet, Vincent; Balian, Gary; Shen, Francis H; Li, Xudong Joshua

2011-06-01

Tissue engineering is a promising approach for treatment of disc degeneration. Herein, we evaluated effects of rotating bioreactor culture on the extracellular matrix production and proliferation of human annulus fibrosus (AF) cells. AF cells were embedded into alginate beads, and then cultured up to 3 weeks in a rotating wall vessel bioreactor or a static vessel. By real-time reverse transcription-polymerase chain reaction, expression of aggrecan, collagen type I and type II, and collagen prolyl 4-hydroxylase II was remarkably elevated, whereas expression of matrix metalloproteinase 3 and a disintegrin and metalloproteinase with thrombospondin motifs 5 was significantly decreased under bioreactor. Biochemical analysis revealed that the levels of the whole cell-associated proteoglycan and collagen were approximately five- and twofolds in rotating bioreactor, respectively, compared to those in static culture. Moreover, AF cell proliferation was augmented in rotating bioreactor. DNA contents were threefolds higher in rotating bioreactor than that in static culture. Expression of the proliferating cell nuclear antigen was robustly enhanced in rotating bioreactor as early as 1 week. Our findings suggested that rotating bioreactor culture would be an effective technique for expansion of human annulus cells for tissue engineering driven treatment of disc degeneration.

Plant cell wall proteomics: the leadership of Arabidopsis thaliana

PubMed Central

Albenne, Cécile; Canut, Hervé; Jamet, Elisabeth

2013-01-01

Plant cell wall proteins (CWPs) progressively emerged as crucial components of cell walls although present in minor amounts. Cell wall polysaccharides such as pectins, hemicelluloses, and cellulose represent more than 90% of primary cell wall mass, whereas hemicelluloses, cellulose, and lignins are the main components of lignified secondary walls. All these polymers provide mechanical properties to cell walls, participate in cell shape and prevent water loss in aerial organs. However, cell walls need to be modified and customized during plant development and in response to environmental cues, thus contributing to plant adaptation. CWPs play essential roles in all these physiological processes and particularly in the dynamics of cell walls, which requires organization and rearrangements of polysaccharides as well as cell-to-cell communication. In the last 10 years, plant cell wall proteomics has greatly contributed to a wider knowledge of CWPs. This update will deal with (i) a survey of plant cell wall proteomics studies with a focus on Arabidopsis thaliana; (ii) the main protein families identified and the still missing peptides; (iii) the persistent issue of the non-canonical CWPs; (iv) the present challenges to overcome technological bottlenecks; and (v) the perspectives beyond cell wall proteomics to understand CWP functions. PMID:23641247

A novel transient wall heat transfer approach for the start-up of SI engines with gasoline direct injection

NASA Astrophysics Data System (ADS)

Lejsek, David; Kulzer, André; Hammer, Jürgen

2010-11-01

The introduction of CO2-reduction technologies like Start-Stop or the Hybrid-Powertrain and the worldwide stringent emission legislation require a detailed optimization of the engine start-up. The combustion concept development as well as the calibration of the engine control unit makes an explicit thermodynamic analysis of the combustion process during the start-up necessary. Initially, the well-known thermodynamic analysis of in-cylinder pressure at stationary condition was transmitted to the highly non-stationary engine start-up. For this running mode of the engine the current models for calculation of the transient wall heat fluxes were found to be misleading. With a fraction of nearly 45% of the burned fuel energy, the wall heat is very important for the calculation of energy balance and for the combustion process analysis. Based on the measurements of transient wall heat transfer densities during the start-up presented in a former work (Lejsek and Kulzer in Investigations on the transient wall heat transfer at start-up for SI engines with gasoline direct injection. SAE Paper), the paper describes the development of adaptations to the known correlations by Woschni (MTZ 31:491, 1970), Hohenberg (Experimentelle Erfassung der Wandwärme von Kolbenmotoren. TU Graz, Habil., 1980) and Bargende (Ein Gleichungsansatz zur Berechnung der instationären Wandwärmeverluste im Hochdruckteil von Ottomotoren. TH Darmstadt, PhD-Thesis, 1991) for the application during engine start-up. To demonstrate the high accuracy of the model, the results of the cyclic resolved thermodynamic analysis using the presented novel approaches were compared with the results of the measurements. It is shown, that the novel heat flux models for the engine start-up process gives a cyclic resolved thermodynamic analysis to optimize the engine start-up pretty efficient.

Cell Wall Structure of Coccoid Green Algae as an Important Trade-Off Between Biotic Interference Mechanisms and Multidimensional Cell Growth.

PubMed

Dunker, Susanne; Wilhelm, Christian

2018-01-01

Coccoid green algae can be divided in two groups based on their cell wall structure. One group has a highly chemical resistant cell wall (HR-cell wall) containing algaenan. The other group is more susceptible to chemicals (LR-cell wall - Low resistant cell wall). Algaenan is considered as important molecule to explain cell wall resistance. Interestingly, cell wall types (LR- and HR-cell wall) are not in accordance with the taxonomic classes Chlorophyceae and Trebouxiophyceae, which makes it even more interesting to consider the ecological function. It was already shown that algaenan helps to protect against virus, bacterial and fungal attack, but in this study we show for the first time that green algae with different cell wall properties show different sensitivity against interference competition with the cyanobacterium Microcystis aeruginosa . Based on previous work with co-cultures of M. aeruginosa and two green algae ( Acutodesmus obliquus and Oocystis marssonii ) differing in their cell wall structure, it was shown that M. aeruginosa could impair only the growth of the green algae if they belong to the LR-cell wall type. In this study it was shown that the sensitivity to biotic interference mechanism shows a more general pattern within coccoid green algae species depending on cell wall structure.

The N-Linked Outer Chain Mannans and the Dfg5p and Dcw1p Endo-α-1,6-Mannanases Are Needed for Incorporation of Candida albicans Glycoproteins into the Cell Wall

PubMed Central

Ao, Jie; Chinnici, Jennifer L.; Maddi, Abhiram

2015-01-01

A biochemical pathway for the incorporation of cell wall protein into the cell wall of Neurospora crassa was recently proposed. In this pathway, the DFG-5 and DCW-1 endo-α-1,6-mannanases function to covalently cross-link cell wall protein-associated N-linked galactomannans, which are structurally related to the yeast outer chain mannans, into the cell wall glucan-chitin matrix. In this report, we demonstrate that the mannosyltransferase enzyme Och1p, which is needed for the synthesis of the N-linked outer chain mannan, is essential for the incorporation of cell wall glycoproteins into the Candida albicans cell wall. Using endoglycosidases, we show that C. albicans cell wall proteins are cross-linked into the cell wall via their N-linked outer chain mannans. We further demonstrate that the Dfg5p and Dcw1p α-1,6-mannanases are needed for the incorporation of cell wall glycoproteins into the C. albicans cell wall. Our results support the hypothesis that the Dfg5p and Dcw1p α-1,6-mannanases incorporate cell wall glycoproteins into the C. albicans cell wall by cross-linking outer chain mannans into the cell wall glucan-chitin matrix. PMID:26048011

Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

PubMed Central

Levin, David E.

2011-01-01

The yeast cell wall is a strong, but elastic, structure that is essential not only for the maintenance of cell shape and integrity, but also for progression through the cell cycle. During growth and morphogenesis, and in response to environmental challenges, the cell wall is remodeled in a highly regulated and polarized manner, a process that is principally under the control of the cell wall integrity (CWI) signaling pathway. This pathway transmits wall stress signals from the cell surface to the Rho1 GTPase, which mobilizes a physiologic response through a variety of effectors. Activation of CWI signaling regulates the production of various carbohydrate polymers of the cell wall, as well as their polarized delivery to the site of cell wall remodeling. This review article centers on CWI signaling in Saccharomyces cerevisiae through the cell cycle and in response to cell wall stress. The interface of this signaling pathway with other pathways that contribute to the maintenance of cell wall integrity is also discussed. PMID:22174182

Study of Periodical Flow Heat Transfer in an Internal Combustion Engine

NASA Astrophysics Data System (ADS)

Luo, Xi

In-cylinder heat transfer is one of the most critical physical behaviors which has a direct influence on engine out emission and thermal efficiency for IC engine. In-cylinder wall temperature has to be precisely controlled to achieve high efficiency and low emission. However, this cannot be done without knowing gas-to-wall heat flux. This study reports on the development of a technique suitable for engine in-cylinder surface temperature measurement, as the traditional method is "hard to reach." A laser induced phosphorescence technique was used to study in-cylinder wall temperature effects on engine out unburned hydrocarbons during the engine transitional period (warm up). A linear correlation was found between the cylinder wall surface temperature and the unburned hydrocarbons at mediate and high charge densities. At low charge density, no clear correlation was observed because of miss-fire events. A new auto background correction infrared (IR) diagnostic was developed to measure the instantaneous in-cylinder surface temperature at 0.1 CAD resolution. A numerical mechanism was designed to suppress relatively low-frequency background noise and provide an accurate in-cylinder surface temperature measurements with an error of less than 1.4% inside the IC engine. In addition, a proposed optical coating reduced time delay errors by 50% compared to more conventional thermocouple techniques. A new cycle-averaged Res number was developed for an IC engine to capture the characteristics of engine flow. Comparison and scaling between different engine flow parameters are available by matching the averaged Res number. From experimental results, the engine flow motion was classified as intermittently turbulent, and it is different from the original fully developed turbulent assumption, which has previously been used in almost all engine simulations. The intermittent turbulence could have a great impact on engine heat transfer because of the transitional turbulence effect. Engine 3D CFD model further proves the existence of transitional turbulence flow. A new multi zone heat transfer model is proposed for IC engines only. The model includes pressure work effects and improved heat transfer prediction compared to the standard Law of the wall model.

4th International Plant Biomechanics Conference Proceedings (Abstracts)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Frank W. Telewski; Lothar H. Koehler; Frank W. Ewers

2003-07-20

The 4th International Plant Biomechanics Conference facilitated an interdisciplinary exchange between scientists, engineers, and educators addressing the major questions encountered in the field of Plant Biomechanics. Subjects covered by the conference include: Evolution; Ecology; Mechanoreception; Cell Walls; Genetic Modification; Applied Biomechanics of Whole Plants, Plant Products, Fibers & Composites; Fluid Dynamics; Wood & Trees; Fracture Mechanics; Xylem Pressure & Water Transport; Modeling; and Introducing Plant Biomechanics in Secondary School Education.

Laser structuring of carbon nanoframe in a protein matrix for the creation of 3D composite materials and coatings for applications in tissue engineering

NASA Astrophysics Data System (ADS)

Gerasimenko, Alexander Yu.; Glukhova, Olga E.; Savostyanov, Georgy V.; Savelyev, Mikhail S.; Ichkitidze, Levan P.; Masloboev, Yurii P.; Selishchev, Sergey V.; Podgaetsky, Vitaly M.

2017-07-01

The results of experimental creation of nanocomposites using femtosecond laser are presented. We have theoretically proved the formation of a carbon nanotube frame in a protein matrix during laser structuring of single-walled carbon nanotubes. We have selected the technological parameters of synthesis of nanocomposites, which provide the proliferation of living cells.

PRECAST CONCRETE WALL PANELS ARE LIFTED INTO PLACE ON MTR ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

PRECAST CONCRETE WALL PANELS ARE LIFTED INTO PLACE ON MTR STEEL FRAME STRUCTURE. INL NEGATIVE NO. 1330. Unknown Photographer, 1/1951 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

FAST CHOPPER BUILDING, TRA665, INTERIOR. UPPER LEVEL. CONCRETE WALLS. INL ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

FAST CHOPPER BUILDING, TRA-665, INTERIOR. UPPER LEVEL. CONCRETE WALLS. INL NEGATIVE NO. HD42-2. Mike Crane, Photographer, 3/2004 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

Numerical simulation of fluid field and in vitro three-dimensional fabrication of tissue-engineered bones in a rotating bioreactor and in vivo implantation for repairing segmental bone defects.

PubMed

Song, Kedong; Wang, Hai; Zhang, Bowen; Lim, Mayasari; Liu, Yingchao; Liu, Tianqing

2013-03-01

In this paper, two-dimensional flow field simulation was conducted to determine shear stresses and velocity profiles for bone tissue engineering in a rotating wall vessel bioreactor (RWVB). In addition, in vitro three-dimensional fabrication of tissue-engineered bones was carried out in optimized bioreactor conditions, and in vivo implantation using fabricated bones was performed for segmental bone defects of Zelanian rabbits. The distribution of dynamic pressure, total pressure, shear stress, and velocity within the culture chamber was calculated for different scaffold locations. According to the simulation results, the dynamic pressure, velocity, and shear stress around the surface of cell-scaffold construction periodically changed at different locations of the RWVB, which could result in periodical stress stimulation for fabricated tissue constructs. However, overall shear stresses were relatively low, and the fluid velocities were uniform in the bioreactor. Our in vitro experiments showed that the number of cells cultured in the RWVB was five times higher than those cultured in a T-flask. The tissue-engineered bones grew very well in the RWVB. This study demonstrates that stress stimulation in an RWVB can be beneficial for cell/bio-derived bone constructs fabricated in an RWVB, with an application for repairing segmental bone defects.

A model for cell wall dissolution in mating yeast cells: polarized secretion and restricted diffusion of cell wall remodeling enzymes induces local dissolution.

PubMed

Huberman, Lori B; Murray, Andrew W

2014-01-01

Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells.

A Model for Cell Wall Dissolution in Mating Yeast Cells: Polarized Secretion and Restricted Diffusion of Cell Wall Remodeling Enzymes Induces Local Dissolution

PubMed Central

Huberman, Lori B.; Murray, Andrew W.

2014-01-01

Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells. PMID:25329559

A study of the native cell wall structures of the marine alga Ventricaria ventricosa (Siphonocladales, Chlorophyceae) using atomic force microscopy.

PubMed

Eslick, Enid M; Beilby, Mary J; Moon, Anthony R

2014-04-01

A substantial proportion of the architecture of the plant cell wall remains unknown with a few cell wall models being proposed. Moreover, even less is known about the green algal cell wall. Techniques that allow direct visualization of the cell wall in as near to its native state are of importance in unravelling the spatial arrangement of cell wall structures and hence in the development of cell wall models. Atomic force microscopy (AFM) was used to image the native cell wall of living cells of Ventricaria ventricosa (V. ventricosa) at high resolution under physiological conditions. The cell wall polymers were identified mainly qualitatively via their structural appearance. The cellulose microfibrils (CMFs) were easily recognizable and the imaging results indicate that the V. ventricosa cell wall has a cross-fibrillar structure throughout. We found the native wall to be abundant in matrix polysaccharides existing in different curing states. The soft phase matrix polysaccharides susceptible by the AFM scanning tip existed as a glutinous fibrillar meshwork, possibly incorporating both the pectic- and hemicellulosic-type substances. The hard phase matrix producing clearer images, revealed coiled fibrillar structures associated with CMFs, sometimes being resolved as globular structures by the AFM tip. The coiling fibrillar structures were also seen in the images of isolated cell wall fragments. The mucilaginous component of the wall was discernible from the gelatinous cell wall matrix as it formed microstructural domains over the surface. AFM has been successful in imaging the native cell wall and revealing novel findings such as the 'coiling fibrillar structures' and cell wall components which have previously not been seen, that is, the gelatinous matrix phase.

Design of a flow perfusion bioreactor system for bone tissue-engineering applications.

PubMed

Bancroft, Gregory N; Sikavitsas, Vassilios I; Mikos, Antonios G

2003-06-01

Several different bioreactors have been investigated for tissue-engineering applications. Among these bioreactors are the spinner flask and the rotating wall vessel reactor. In addition, a new type of culture system has been developed and investigated, the flow perfusion culture bioreactor. Flow perfusion culture offers several advantages, notably the ability to mitigate both external and internal diffusional limitations as well as to apply mechanical stress to the cultured cells. For such investigation, a flow perfusion culture system was designed and built. This design is the outgrowth of important design requirements and incorporates features crucial to successful experimentation with such a system.

Altered Cell Wall Plasticity Can Restrict Plant Growth under Ammonium Nutrition.

PubMed

Podgórska, Anna; Burian, Maria; Gieczewska, Katarzyna; Ostaszewska-Bugajska, Monika; Zebrowski, Jacek; Solecka, Danuta; Szal, Bożena

2017-01-01

Plants mainly utilize inorganic forms of nitrogen (N), such as nitrate (NO 3 - ) and ammonium (NH 4 + ). However, the composition of the N source is important, because excess of NH 4 + promotes morphological disorders. Plants cultured on NH 4 + as the sole N source exhibit serious growth inhibition, commonly referred to as "ammonium toxicity syndrome." NH 4 + -mediated suppression of growth may be attributable to both repression of cell elongation and reduction of cell division. The precondition for cell enlargement is the expansion of the cell wall, which requires the loosening of the cell wall polymers. Therefore, to understand how NH 4 + nutrition may trigger growth retardation in plants, properties of their cell walls were analyzed. We found that Arabidopsis thaliana using NH 4 + as the sole N source has smaller cells with relatively thicker cell walls. Moreover, cellulose, which is the main load-bearing polysaccharide revealed a denser assembly of microfibrils. Consequently, the leaf blade tissue showed elevated tensile strength and indicated higher cell wall stiffness. These changes might be related to changes in polysaccharide and ion content of cell walls. Further, NH 4 + toxicity was associated with altered activities of cell wall modifying proteins. The lower activity and/or expression of pectin hydrolyzing enzymes and expansins might limit cell wall expansion. Additionally, the higher activity of cell wall peroxidases can lead to higher cross-linking of cell wall polymers. Overall, the NH 4 + -mediated inhibition of growth is related to a more rigid cell wall structure, which limits expansion of cells. The changes in cell wall composition were also indicated by decreased expression of Feronia , a receptor-like kinase involved in the control of cell wall extension.

Altered Cell Wall Plasticity Can Restrict Plant Growth under Ammonium Nutrition

PubMed Central

Podgórska, Anna; Burian, Maria; Gieczewska, Katarzyna; Ostaszewska-Bugajska, Monika; Zebrowski, Jacek; Solecka, Danuta; Szal, Bożena

2017-01-01

Plants mainly utilize inorganic forms of nitrogen (N), such as nitrate (NO3–) and ammonium (NH4+). However, the composition of the N source is important, because excess of NH4+ promotes morphological disorders. Plants cultured on NH4+ as the sole N source exhibit serious growth inhibition, commonly referred to as “ammonium toxicity syndrome.” NH4+-mediated suppression of growth may be attributable to both repression of cell elongation and reduction of cell division. The precondition for cell enlargement is the expansion of the cell wall, which requires the loosening of the cell wall polymers. Therefore, to understand how NH4+ nutrition may trigger growth retardation in plants, properties of their cell walls were analyzed. We found that Arabidopsis thaliana using NH4+ as the sole N source has smaller cells with relatively thicker cell walls. Moreover, cellulose, which is the main load-bearing polysaccharide revealed a denser assembly of microfibrils. Consequently, the leaf blade tissue showed elevated tensile strength and indicated higher cell wall stiffness. These changes might be related to changes in polysaccharide and ion content of cell walls. Further, NH4+ toxicity was associated with altered activities of cell wall modifying proteins. The lower activity and/or expression of pectin hydrolyzing enzymes and expansins might limit cell wall expansion. Additionally, the higher activity of cell wall peroxidases can lead to higher cross-linking of cell wall polymers. Overall, the NH4+-mediated inhibition of growth is related to a more rigid cell wall structure, which limits expansion of cells. The changes in cell wall composition were also indicated by decreased expression of Feronia, a receptor-like kinase involved in the control of cell wall extension. PMID:28848567

Lipid engineering reveals regulatory roles for membrane fluidity in yeast flocculation and oxygen-limited growth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Degreif, Daniel; de Rond, Tristan; Bertl, Adam

Cells modulate lipid metabolism in order to maintain membrane homeostasis. In this paper, we use a metabolic engineering approach to manipulate the stoichiometry of fatty acid unsaturation, a regulator of cell membrane fluidity, in Saccharomyces cerevisiae. Unexpectedly, reduced lipid unsaturation triggered cell-cell adhesion (flocculation), a phenomenon characteristic of industrial yeast but uncommon in laboratory strains. We find that ER lipid saturation sensors induce expression of FLO1 – encoding a cell wall polysaccharide binding protein – independently of its canonical regulator. In wild-type cells, Flo1p-dependent flocculation occurs under oxygen-limited growth, which reduces unsaturated lipid synthesis and thus serves as the environmentalmore » trigger for flocculation. Transcriptional analysis shows that FLO1 is one of the most highly induced genes in response to changes in lipid unsaturation, and that the set of membrane fluidity-sensitive genes is globally activated as part of the cell's long-term response to hypoxia during fermentation. Finally, our results show how the lipid homeostasis machinery of budding yeast is adapted to carry out a broad response to an environmental stimulus important in biotechnology.« less

Label-free in situ Imaging of Lignification in Plant Cell Walls

PubMed Central

Schmidt, Martin; Perera, Pradeep; Schwartzberg, Adam M.; Adams, Paul D.; Schuck, P. James

2010-01-01

Meeting growing energy demands safely and efficiently is a pressing global challenge. Therefore, research into biofuels production that seeks to find cost-effective and sustainable solutions has become a topical and critical task. Lignocellulosic biomass is poised to become the primary source of biomass for the conversion to liquid biofuels1-6. However, the recalcitrance of these plant cell wall materials to cost-effective and efficient degradation presents a major impediment for their use in the production of biofuels and chemicals4. In particular, lignin, a complex and irregular poly-phenylpropanoid heteropolymer, becomes problematic to the postharvest deconstruction of lignocellulosic biomass. For example in biomass conversion for biofuels, it inhibits saccharification in processes aimed at producing simple sugars for fermentation7. The effective use of plant biomass for industrial purposes is in fact largely dependent on the extent to which the plant cell wall is lignified. The removal of lignin is a costly and limiting factor8 and lignin has therefore become a key plant breeding and genetic engineering target in order to improve cell wall conversion. Analytical tools that permit the accurate rapid characterization of lignification of plant cell walls become increasingly important for evaluating a large number of breeding populations. Extractive procedures for the isolation of native components such as lignin are inevitably destructive, bringing about significant chemical and structural modifications9-11. Analytical chemical in situ methods are thus invaluable tools for the compositional and structural characterization of lignocellulosic materials. Raman microscopy is a technique that relies on inelastic or Raman scattering of monochromatic light, like that from a laser, where the shift in energy of the laser photons is related to molecular vibrations and presents an intrinsic label-free molecular "fingerprint" of the sample. Raman microscopy can afford non-destructive and comparatively inexpensive measurements with minimal sample preparation, giving insights into chemical composition and molecular structure in a close to native state. Chemical imaging by confocal Raman microscopy has been previously used for the visualization of the spatial distribution of cellulose and lignin in wood cell walls12-14. Based on these earlier results, we have recently adopted this method to compare lignification in wild type and lignin-deficient transgenic Populus trichocarpa (black cottonwood) stem wood15. Analyzing the lignin Raman bands16,17 in the spectral region between 1,600 and 1,700 cm-1, lignin signal intensity and localization were mapped in situ. Our approach visualized differences in lignin content, localization, and chemical composition. Most recently, we demonstrated Raman imaging of cell wall polymers in Arabidopsis thaliana with lateral resolution that is sub-μm18. Here, this method is presented affording visualization of lignin in plant cell walls and comparison of lignification in different tissues, samples or species without staining or labeling of the tissues. PMID:21085100

Tissue-engineered microenvironment systems for modeling human vasculature.

PubMed

Tourovskaia, Anna; Fauver, Mark; Kramer, Gregory; Simonson, Sara; Neumann, Thomas

2014-09-01

The high attrition rate of drug candidates late in the development process has led to an increasing demand for test assays that predict clinical outcome better than conventional 2D cell culture systems and animal models. Government agencies, the military, and the pharmaceutical industry have started initiatives for the development of novel in-vitro systems that recapitulate functional units of human tissues and organs. There is growing evidence that 3D cell arrangement, co-culture of different cell types, and physico-chemical cues lead to improved predictive power. A key element of all tissue microenvironments is the vasculature. Beyond transporting blood the microvasculature assumes important organ-specific functions. It is also involved in pathologic conditions, such as inflammation, tumor growth, metastasis, and degenerative diseases. To provide a tool for modeling this important feature of human tissue microenvironments, we developed a microfluidic chip for creating tissue-engineered microenvironment systems (TEMS) composed of tubular cell structures. Our chip design encompasses a small chamber that is filled with an extracellular matrix (ECM) surrounding one or more tubular channels. Endothelial cells (ECs) seeded into the channels adhere to the ECM walls and grow into perfusable tubular tissue structures that are fluidically connected to upstream and downstream fluid channels in the chip. Using these chips we created models of angiogenesis, the blood-brain barrier (BBB), and tumor-cell extravasation. Our angiogenesis model recapitulates true angiogenesis, in which sprouting occurs from a "parent" vessel in response to a gradient of growth factors. Our BBB model is composed of a microvessel generated from brain-specific ECs within an ECM populated with astrocytes and pericytes. Our tumor-cell extravasation model can be utilized to visualize and measure tumor-cell migration through vessel walls into the surrounding matrix. The described technology can be used to create TEMS that recapitulate structural, functional, and physico-chemical elements of vascularized human tissue microenvironments in vitro. © 2014 by the Society for Experimental Biology and Medicine.

Experimental investigations of heat transfer and temperature fields in models simulating fuel assemblies used in the core of a nuclear reactor with a liquid heavy-metal coolant

NASA Astrophysics Data System (ADS)

Belyaev, I. A.; Genin, L. G.; Krylov, S. G.; Novikov, A. O.; Razuvanov, N. G.; Sviridov, V. G.

2015-09-01

The aim of this experimental investigation is to obtain information on the temperature fields and heat transfer coefficients during flow of liquid-metal coolant in models simulating an elementary cell in the core of a liquid heavy metal cooled fast-neutron reactor. Two design versions for spacing fuel rods in the reactor core were considered. In the first version, the fuel rods were spaced apart from one another using helical wire wound on the fuel rod external surface, and in the second version spacer grids were used for the same purpose. The experiments were carried out on the mercury loop available at the Moscow Power Engineering Institute National Research University's Chair of Engineering Thermal Physics. Two experimental sections simulating an elementary cell for each of the fuel rod spacing versions were fabricated. The temperature fields were investigated using a dedicated hinged probe that allows temperature to be measured at any point of the studied channel cross section. The heat-transfer coefficients were determined using the wall temperature values obtained at the moment when the probe thermocouple tail end touched the channel wall. Such method of determining the wall temperature makes it possible to alleviate errors that are unavoidable in case of measuring the wall temperature using thermocouples placed in slots milled in the wall. In carrying out the experiments, an automated system of scientific research was applied, which allows a large body of data to be obtained within a short period of time. The experimental investigations in the first test section were carried out at Re = 8700, and in the second one, at five values of Reynolds number. Information about temperature fields was obtained by statistically processing the array of sampled probe thermocouple indications at 300 points in the experimental channel cross section. Reach material has been obtained for verifying the codes used for calculating velocity and temperature fields in channels with an intricately shaped cross section simulating the flow pass sections for liquid-metal coolants cooling the core of nuclear reactors.

The Cell Wall of the Human Fungal Pathogen Aspergillus fumigatus: Biosynthesis, Organization, Immune Response, and Virulence.

PubMed

Latgé, Jean-Paul; Beauvais, Anne; Chamilos, Georgios

2017-09-08

More than 90% of the cell wall of the filamentous fungus Aspergillus fumigatus comprises polysaccharides. Biosynthesis of the cell wall polysaccharides is under the control of three types of enzymes: transmembrane synthases, which are anchored to the plasma membrane and use nucleotide sugars as substrates, and cell wall-associated transglycosidases and glycosyl hydrolases, which are responsible for remodeling the de novo synthesized polysaccharides and establishing the three-dimensional structure of the cell wall. For years, the cell wall was considered an inert exoskeleton of the fungal cell. The cell wall is now recognized as a living organelle, since the composition and cellular localization of the different constitutive cell wall components (especially of the outer layers) vary when the fungus senses changes in the external environment. The cell wall plays a major role during infection. The recognition of the fungal cell wall by the host is essential in the initiation of the immune response. The interactions between the different pattern-recognition receptors (PRRs) and cell wall pathogen-associated molecular patterns (PAMPs) orientate the host response toward either fungal death or growth, which would then lead to disease development. Understanding the molecular determinants of the interplay between the cell wall and host immunity is fundamental to combatting Aspergillus diseases.

Cell Wall Composition and Candidate Biosynthesis Gene Expression During Rice Development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Fan; Manisseri, Chithra; Fagerström, Alexandra

Cell walls of grasses, including cereal crops and biofuel grasses, comprise the majority of plant biomass and intimately influence plant growth, development and physiology. However, the functions of many cell wall synthesis genes, and the relationships among and the functions of cell wall components remain obscure. To better understand the patterns of cell wall accumulation and identify genes that act in grass cell wall biosynthesis, we characterized 30 samples from aerial organs of rice (Oryza sativa cv. Kitaake) at 10 developmental time points, 3-100 d post-germination. Within these samples, we measured 15 cell wall chemical components, enzymatic digestibility and 18more » cell wall polysaccharide epitopes/ligands. We also used quantitative reverse transcription-PCR to measure expression of 50 glycosyltransferases, 15 acyltransferases and eight phenylpropanoid genes, many of which had previously been identified as being highly expressed in rice. Most cell wall components vary significantly during development, and correlations among them support current understanding of cell walls. We identified 92 significant correlations between cell wall components and gene expression and establish nine strong hypotheses for genes that synthesize xylans, mixed linkage glucan and pectin components. This work provides an extensive analysis of cell wall composition throughout rice development, identifies genes likely to synthesize grass cell walls, and provides a framework for development of genetically improved grasses for use in lignocellulosic biofuel production and agriculture.« less

An Engineered Monolignol 4-O-Methyltransferase Depresses Lignin Biosynthesis and Confers Novel Metabolic Capability in Arabidopsis[C][W][OA

PubMed Central

Zhang, Kewei; Bhuiya, Mohammad-Wadud; Pazo, Jorge Rencoret; Miao, Yuchen; Kim, Hoon; Ralph, John; Liu, Chang-Jun

2012-01-01

Although the practice of protein engineering is industrially fruitful in creating biocatalysts and therapeutic proteins, applications of analogous techniques in the field of plant metabolic engineering are still in their infancy. Lignins are aromatic natural polymers derived from the oxidative polymerization of primarily three different hydroxycinnamyl alcohols, the monolignols. Polymerization of lignin starts with the oxidation of monolignols, followed by endwise cross-coupling of (radicals of) a monolignol and the growing oligomer/polymer. The para-hydroxyl of each monolignol is crucial for radical generation and subsequent coupling. Here, we describe the structure-function analysis and catalytic improvement of an artificial monolignol 4-O-methyltransferase created by iterative saturation mutagenesis and its use in modulating lignin and phenylpropanoid biosynthesis. We show that expressing the created enzyme in planta, thus etherifying the para-hydroxyls of lignin monomeric precursors, denies the derived monolignols any participation in the subsequent coupling process, substantially reducing lignification and, ultimately, lignin content. Concomitantly, the transgenic plants accumulated de novo synthesized 4-O-methylated soluble phenolics and wall-bound esters. The lower lignin levels of transgenic plants resulted in higher saccharification yields. Our study, through a structure-based protein engineering approach, offers a novel strategy for modulating phenylpropanoid/lignin biosynthesis to improve cell wall digestibility and diversify the repertories of biologically active compounds. PMID:22851762

The Impact of Microfibril Orientations on the Biomechanics of Plant Cell Walls and Tissues.

PubMed

Ptashnyk, Mariya; Seguin, Brian

2016-11-01

The microscopic structure and anisotropy of plant cell walls greatly influence the mechanical properties, morphogenesis, and growth of plant cells and tissues. The microscopic structure and properties of cell walls are determined by the orientation and mechanical properties of the cellulose microfibrils and the mechanical properties of the cell wall matrix. Viewing the shape of a plant cell as a square prism with the axis aligning with the primary direction of expansion and growth, the orientation of the microfibrils within the side walls, i.e. the parts of the cell walls on the sides of the cells, is known. However, not much is known about their orientation at the upper and lower ends of the cell. Here we investigate the impact of the orientation of cellulose microfibrils within the upper and lower parts of the plant cell walls by solving the equations of linear elasticity numerically. Three different scenarios for the orientation of the microfibrils are considered. We also distinguish between the microstructure in the side walls given by microfibrils perpendicular to the main direction of the expansion and the situation where the microfibrils are rotated through the wall thickness. The macroscopic elastic properties of the cell wall are obtained using homogenization theory from the microscopic description of the elastic properties of the cell wall microfibrils and wall matrix. It is found that the orientation of the microfibrils in the upper and lower parts of the cell walls affects the expansion of the cell in the lateral directions and is particularly important in the case of forces acting on plant cell walls and tissues.

High temperature induced disruption of the cell wall integrity and structure in Pleurotus ostreatus mycelia.

PubMed

Qiu, Zhiheng; Wu, Xiangli; Gao, Wei; Zhang, Jinxia; Huang, Chenyang

2018-05-30

Fungal cells are surrounded by a tight cell wall to protect them from harmful environmental conditions and to resist lysis. The synthesis and assembly determine the shape, structure, and integrity of the cell wall during the process of mycelial growth and development. High temperature is an important abiotic stress, which affects the synthesis and assembly of cell walls. In the present study, the chitin and β-1,3-glucan concentrations in the cell wall of Pleurotus ostreatus mycelia were changed after high-temperature treatment. Significantly higher chitin and β-1,3-glucan concentrations were detected at 36 °C than those incubated at 28 °C. With the increased temperature, many aberrant chitin deposition patches occurred, and the distribution of chitin in the cell wall was uneven. Moreover, high temperature disrupts the cell wall integrity, and P. ostreatus mycelia became hypersensitive to cell wall-perturbing agents at 36 °C. The cell wall structure tended to shrink or distorted after high temperature. The cell walls were observed to be thicker and looser by using transmission electron microscopy. High temperature can decrease the mannose content in the cell wall and increase the relative cell wall porosity. According to infrared absorption spectrum, high temperature broke or decreased the glycosidic linkages. Finally, P. ostreatus mycelial cell wall was easily degraded by lysing enzymes after high-temperature treatment. In other words, the cell wall destruction caused by high temperature may be a breakthrough for P. ostreatus to be easily infected by Trichoderma.

Endoplasmic reticulum-derived reactive oxygen species (ROS) is involved in toxicity of cell wall stress to Candida albicans.

PubMed

Yu, Qilin; Zhang, Bing; Li, Jianrong; Zhang, Biao; Wang, Honggang; Li, Mingchun

2016-10-01

The cell wall is an important cell structure in both fungi and bacteria, and hence becomes a common antimicrobial target. The cell wall-perturbing agents disrupt synthesis and function of cell wall components, leading to cell wall stress and consequent cell death. However, little is known about the detailed mechanisms by which cell wall stress renders fungal cell death. In this study, we found that ROS scavengers drastically attenuated the antifungal effect of cell wall-perturbing agents to the model fungal pathogen Candida albicans, and these agents caused remarkable ROS accumulation and activation of oxidative stress response (OSR) in this fungus. Interestingly, cell wall stress did not cause mitochondrial dysfunction and elevation of mitochondrial superoxide levels. Furthermore, the iron chelator 2,2'-bipyridyl (BIP) and the hydroxyl radical scavengers could not attenuate cell wall stress-caused growth inhibition and ROS accumulation. However, cell wall stress up-regulated expression of unfold protein response (UPR) genes, enhanced protein secretion and promoted protein folding-related oxidation of Ero1, an important source of ROS production. These results indicated that oxidation of Ero1 in the endoplasmic reticulum (ER), rather than mitochondrial electron transport and Fenton reaction, contributed to cell wall stress-related ROS accumulation and consequent growth inhibition. Our findings uncover a novel link between cell wall integrity (CWI), ER function and ROS production in fungal cells, and shed novel light on development of strategies promoting the antifungal efficacy of cell wall-perturbing agents against fungal infections. Copyright © 2016 Elsevier Inc. All rights reserved.

Regulation of cell wall biosynthesis.

PubMed

Zhong, Ruiqin; Ye, Zheng-Hua

2007-12-01

Plant cell walls differ in their amount and composition among various cell types and even in different microdomains of the wall of a given cell. Plants must have evolved regulatory mechanisms controlling biosynthesis, targeted secretion, and assembly of wall components to achieve the heterogeneity in cell walls. A number of factors, including hormones, the cytoskeleton, glycosylphosphatidylinositol-anchored proteins, phosphoinositides, and sugar nucleotide supply, have been implicated in the regulation of cell wall biosynthesis or deposition. In the past two years, there have been important discoveries in transcriptional regulation of secondary wall biosynthesis. Several transcription factors in the NAC and MYB families have been shown to be the key switches for activation of secondary wall biosynthesis. These studies suggest a transcriptional network comprised of a hierarchy of transcription factors is involved in regulating secondary wall biosynthesis. Further investigation and integration of the regulatory players participating in the making of cell walls will certainly lead to our understanding of how wall amounts and composition are controlled in a given cell type. This may eventually allow custom design of plant cell walls on the basis of our needs.

Relating Nanoscale Accessibility within Plant Cell Walls to Improved Enzyme Hydrolysis Yields in Corn Stover Subjected to Diverse Pretreatments.

PubMed

Crowe, Jacob D; Zarger, Rachael A; Hodge, David B

2017-10-04

Simultaneous chemical modification and physical reorganization of plant cell walls via alkaline hydrogen peroxide or liquid hot water pretreatment can alter cell wall structural properties impacting nanoscale porosity. Nanoscale porosity was characterized using solute exclusion to assess accessible pore volumes, water retention value as a proxy for accessible water-cell walls surface area, and solute-induced cell wall swelling to measure cell wall rigidity. Key findings concluded that delignification by alkaline hydrogen peroxide pretreatment decreased cell wall rigidity and that the subsequent cell wall swelling resulted increased nanoscale porosity and improved enzyme binding and hydrolysis compared to limited swelling and increased accessible surface areas observed in liquid hot water pretreated biomass. The volume accessible to a 90 Å dextran probe within the cell wall was found to be correlated to both enzyme binding and glucose hydrolysis yields, indicating cell wall porosity is a key contributor to effective hydrolysis yields.

Following the compositional changes of fresh grape skin cell walls during the fermentation process in the presence and absence of maceration enzymes.

PubMed

Zietsman, Anscha J J; Moore, John P; Fangel, Jonatan U; Willats, William G T; Trygg, Johan; Vivier, Melané A

2015-03-18

Cell wall profiling technologies were used to follow compositional changes that occurred in the skins of grape berries (from two different ripeness levels) during fermentation and enzyme maceration. Multivariate data analysis showed that the fermentation process yielded cell walls enriched in hemicellulose components because pectin was solubilized (and removed) with a reduction as well as exposure of cell wall proteins usually embedded within the cell wall structure. The addition of enzymes caused even more depectination, and the enzymes unravelled the cell walls enabling better access to, and extraction of, all cell wall polymers. Overripe grapes had cell walls that were extensively hydrolyzed and depolymerized, probably by natural grape-tissue-ripening enzymes, and this enhanced the impact that the maceration enzymes had on the cell wall monosaccharide profile. The combination of the techniques that were used is an effective direct measurement of the hydrolysis actions of maceration enzymes on the cell walls of grape berry skin.

Employing proteomic analysis to compare Paracoccidioides lutzii yeast and mycelium cell wall proteins.

PubMed

Araújo, Danielle Silva; de Sousa Lima, Patrícia; Baeza, Lilian Cristiane; Parente, Ana Flávia Alves; Melo Bailão, Alexandre; Borges, Clayton Luiz; de Almeida Soares, Célia Maria

2017-11-01

Paracoccidioidomycosis is an important systemic mycosis caused by thermodimorphic fungi of the Paracoccidioides genus. During the infective process, the cell wall acts at the interface between the fungus and the host. In this way, the cell wall has a key role in growth, environment sensing and interaction, as well as morphogenesis of the fungus. Since the cell wall is absent in mammals, it may present molecules that are described as target sites for new antifungal drugs. Despite its importance, up to now few studies have been conducted employing proteomics in for the identification of cell wall proteins in Paracoccidioides spp. Here, a detailed proteomic approach, including cell wall-fractionation coupled to NanoUPLC-MS E , was used to study and compare the cell wall fractions from Paracoccidioides lutzii mycelia and yeast cells. The analyzed samples consisted of cell wall proteins extracted by hot SDS followed by extraction by mild alkali. In summary, 512 proteins constituting different cell wall fractions were identified, including 7 predicted GPI-dependent cell wall proteins that are potentially involved in cell wall metabolism. Adhesins previously described in Paracoccidioides spp. such as enolase, glyceraldehyde-3-phosphate dehydrogenase were identified. Comparing the proteins in mycelium and yeast cells, we detected some that are common to both fungal phases, such as Ecm33, and some specific proteins, as glucanase Crf1. All of those proteins were described in the metabolism of cell wall. Our study provides an important elucidation of cell wall composition of fractions in Paracoccidioides, opening a way to understand the fungus cell wall architecture. Copyright © 2017 Elsevier B.V. All rights reserved.

Fabrication of cell container arrays with overlaid surface topographies.

PubMed

Truckenmüller, Roman; Giselbrecht, Stefan; Escalante-Marun, Maryana; Groenendijk, Max; Papenburg, Bernke; Rivron, Nicolas; Unadkat, Hemant; Saile, Volker; Subramaniam, Vinod; van den Berg, Albert; van Blitterswijk, Clemens; Wessling, Matthias; de Boer, Jan; Stamatialis, Dimitrios

2012-02-01

This paper presents cell culture substrates in the form of microcontainer arrays with overlaid surface topographies, and a technology for their fabrication. The new fabrication technology is based on microscale thermoforming of thin polymer films whose surfaces are topographically prepatterned on a micro- or nanoscale. For microthermoforming, we apply a new process on the basis of temporary back moulding of polymer films and use the novel concept of a perforated-sheet-like mould. Thermal micro- or nanoimprinting is applied for prepatterning. The novel cell container arrays are fabricated from polylactic acid (PLA) films. The thin-walled microcontainer structures have the shape of a spherical calotte merging into a hexagonal shape at their upper circumferential edges. In the arrays, the cell containers are arranged densely packed in honeycomb fashion. The inner surfaces of the highly curved container walls are provided with various topographical micro- and nanopatterns. For a first validation of the microcontainer arrays as in vitro cell culture substrates, C2C12 mouse premyoblasts are cultured in containers with microgrooved surfaces and shown to align along the grooves in the three-dimensional film substrates. In future stem-cell-biological and tissue engineering applications, microcontainers fabricated using the proposed technology may act as geometrically defined artificial microenvironments or niches.

Salt stress causes cell wall damage in yeast cells lacking mitochondrial DNA.

PubMed

Gao, Qiuqiang; Liou, Liang-Chun; Ren, Qun; Bao, Xiaoming; Zhang, Zhaojie

2014-03-03

The yeast cell wall plays an important role in maintaining cell morphology, cell integrity and response to environmental stresses. Here, we report that salt stress causes cell wall damage in yeast cells lacking mitochondrial DNA (ρ 0 ). Upon salt treatment, the cell wall is thickened, broken and becomes more sensitive to the cell wall-perturbing agent sodium dodecyl sulfate (SDS). Also, SCW11 mRNA levels are elevated in ρ 0 cells. Deletion of SCW11 significantly decreases the sensitivity of ρ 0 cells to SDS after salt treatment, while overexpression of SCW11 results in higher sensitivity. In addition, salt stress in ρ 0 cells induces high levels of reactive oxygen species (ROS), which further damages the cell wall, causing cells to become more sensitive towards the cell wall-perturbing agent.

Non-preconditioned conjugate gradient on cell and FPGA based hybrid supercomputer nodes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dubois, David H; Dubois, Andrew J; Boorman, Thomas M

2009-01-01

This work presents a detailed implementation of a double precision, non-preconditioned, Conjugate Gradient algorithm on a Roadrunner heterogeneous supercomputer node. These nodes utilize the Cell Broadband Engine Architecture{sup TM} in conjunction with x86 Opteron{sup TM} processors from AMD. We implement a common Conjugate Gradient algorithm, on a variety of systems, to compare and contrast performance. Implementation results are presented for the Roadrunner hybrid supercomputer, SRC Computers, Inc. MAPStation SRC-6 FPGA enhanced hybrid supercomputer, and AMD Opteron only. In all hybrid implementations wall clock time is measured, including all transfer overhead and compute timings.

Non-preconditioned conjugate gradient on cell and FPCA-based hybrid supercomputer nodes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dubois, David H; Dubois, Andrew J; Boorman, Thomas M

2009-03-10

This work presents a detailed implementation of a double precision, Non-Preconditioned, Conjugate Gradient algorithm on a Roadrunner heterogeneous supercomputer node. These nodes utilize the Cell Broadband Engine Architecture{trademark} in conjunction with x86 Opteron{trademark} processors from AMD. We implement a common Conjugate Gradient algorithm, on a variety of systems, to compare and contrast performance. Implementation results are presented for the Roadrunner hybrid supercomputer, SRC Computers, Inc. MAPStation SRC-6 FPGA enhanced hybrid supercomputer, and AMD Opteron only. In all hybrid implementations wall clock time is measured, including all transfer overhead and compute timings.

Engineering a monolignol 4-O-methyltransferase with high selectivity for the condensed lignin precursor coniferyl alcohol.

PubMed

Cai, Yuanheng; Bhuiya, Mohammad-Wadud; Shanklin, John; Liu, Chang-Jun

2015-10-30

Lignin, a rigid biopolymer in plant cell walls, is derived from the oxidative polymerization of three monolignols. The composition of monolignol monomers dictates the degree of lignin condensation, reactivity, and thus the degradability of plant cell walls. Guaiacyl lignin is regarded as the condensed structural unit. Polymerization of lignin is initiated through the deprotonation of the para-hydroxyl group of monolignols. Therefore, preferentially modifying the para-hydroxyl of a specific monolignol to deprive its dehydrogenation propensity would disturb the formation of particular lignin subunits. Here, we test the hypothesis that specific remodeling the active site of a monolignol 4-O-methyltransferase would create an enzyme that specifically methylates the condensed guaiacyl lignin precursor coniferyl alcohol. Combining crystal structural information with combinatorial active site saturation mutagenesis and starting with the engineered promiscuous enzyme, MOMT5 (T133L/E165I/F175I/F166W/H169F), we incrementally remodeled its substrate binding pocket by the addition of four substitutions, i.e. M26H, S30R, V33S, and T319M, yielding a mutant enzyme capable of discriminately etherifying the para-hydroxyl of coniferyl alcohol even in the presence of excess sinapyl alcohol. The engineered enzyme variant has a substantially reduced substrate binding pocket that imposes a clear steric hindrance thereby excluding bulkier lignin precursors. The resulting enzyme variant represents an excellent candidate for modulating lignin composition and/or structure in planta. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Pectic homogalacturonan masks abundant sets of xyloglucan epitopes in plant cell walls.

PubMed

Marcus, Susan E; Verhertbruggen, Yves; Hervé, Cécile; Ordaz-Ortiz, José J; Farkas, Vladimir; Pedersen, Henriette L; Willats, William G T; Knox, J Paul

2008-05-22

Molecular probes are required to detect cell wall polymers in-situ to aid understanding of their cell biology and several studies have shown that cell wall epitopes have restricted occurrences across sections of plant organs indicating that cell wall structure is highly developmentally regulated. Xyloglucan is the major hemicellulose or cross-linking glycan of the primary cell walls of dicotyledons although little is known of its occurrence or functions in relation to cell development and cell wall microstructure. Using a neoglycoprotein approach, in which a XXXG heptasaccharide of tamarind seed xyloglucan was coupled to BSA to produce an immunogen, we have generated a rat monoclonal antibody (designated LM15) to the XXXG structural motif of xyloglucans. The specificity of LM15 has been confirmed by the analysis of LM15 binding using glycan microarrays and oligosaccharide hapten inhibition of binding studies. The use of LM15 for the analysis of xyloglucan in the cell walls of tamarind and nasturtium seeds, in which xyloglucan occurs as a storage polysaccharide, indicated that the LM15 xyloglucan epitope occurs throughout the thickened cell walls of the tamarind seed and in the outer regions, adjacent to middle lamellae, of the thickened cell walls of the nasturtium seed. Immunofluorescence analysis of LM15 binding to sections of tobacco and pea stem internodes indicated that the xyloglucan epitope was restricted to a few cell types in these organs. Enzymatic removal of pectic homogalacturonan from equivalent sections resulted in the abundant detection of distinct patterns of the LM15 xyloglucan epitope across these organs and a diversity of occurrences in relation to the cell wall microstructure of a range of cell types. These observations support ideas that xyloglucan is associated with pectin in plant cell walls. They also indicate that documented patterns of cell wall epitopes in relation to cell development and cell differentiation may need to be re-considered in relation to the potential masking of cell wall epitopes by other cell wall components.

The Interplay between Cell Wall Mechanical Properties and the Cell Cycle in Staphylococcus aureus

PubMed Central

Bailey, Richard G.; Turner, Robert D.; Mullin, Nic; Clarke, Nigel; Foster, Simon J.; Hobbs, Jamie K.

2014-01-01

The nanoscale mechanical properties of live Staphylococcus aureus cells during different phases of growth were studied by atomic force microscopy. Indentation to different depths provided access to both local cell wall mechanical properties and whole-cell properties, including a component related to cell turgor pressure. Local cell wall properties were found to change in a characteristic manner throughout the division cycle. Splitting of the cell into two daughter cells followed a local softening of the cell wall along the division circumference, with the cell wall on either side of the division circumference becoming stiffer. Once exposed, the newly formed septum was found to be stiffer than the surrounding, older cell wall. Deeper indentations, which were affected by cell turgor pressure, did not show a change in stiffness throughout the division cycle, implying that enzymatic cell wall remodeling and local variations in wall properties are responsible for the evolution of cell shape through division. PMID:25468333

Antisense expression of the fasciclin-like arabinogalactan protein FLA6 gene in Populus inhibits expression of its homologous genes and alters stem biomechanics and cell wall composition in transgenic trees

PubMed Central

Wang, Haihai; Jiang, Chunmei; Wang, Cuiting; Yang, Yang; Yang, Lei; Gao, Xiaoyan; Zhang, Hongxia

2015-01-01

Fasciclin-like arabinogalactan proteins (FLAs) play important roles in the growth and development of roots, stems, and seeds in Arabidopsis. However, their biological functions in woody plants are largely unknown. In this work, we investigated the possible function of PtFLA6 in poplar. Quantitative real-time PCR, PtFLA6–yellow fluorescent protein (YFP) fusion protein subcellular localization, Western blotting, and immunohistochemical analyses demonstrated that the PtFLA6 gene was expressed specifically in the xylem of mature stem, and PtFLA6 protein was distributed ubiquitous in plant cells and accumulated predominantly in stem xylem fibres. Antisense expression of PtFLA6 in the aspen hybrid clone Poplar davidiana×Poplar bolleana reduced the transcripts of PtFLA6 and its homologous genes. Transgenic plants that showed a significant reduction in the transcripts of PtFLAs accumulated fewer PtFLA6 and arabinogalactan proteins than did the non-transgenic plants, leading to reduced stem flexural strength and stiffness. Further studies revealed that the altered stem biomechanics of transgenic plants could be attributed to the decreased cellulose and lignin composition in the xylem. In addition expression of some xylem-specific genes involved in cell wall biosynthesis was downregulated in these transgenic plants. All these results suggest that engineering the expression of PtFLA6 and its homologues could modulate stem mechanical properties by affecting cell wall composition in trees. PMID:25428999

KRE5 Suppression Induces Cell Wall Stress and Alternative ER Stress Response Required for Maintaining Cell Wall Integrity in Candida glabrata

PubMed Central

Sasaki, Masato; Ito, Fumie; Aoyama, Toshio; Sato-Okamoto, Michiyo; Takahashi-Nakaguchi, Azusa; Chibana, Hiroji; Shibata, Nobuyuki

2016-01-01

The maintenance of cell wall integrity in fungi is required for normal cell growth, division, hyphae formation, and antifungal tolerance. We observed that endoplasmic reticulum stress regulated cell wall integrity in Candida glabrata, which possesses uniquely evolved mechanisms for unfolded protein response mechanisms. Tetracycline-mediated suppression of KRE5, which encodes a predicted UDP-glucose:glycoprotein glucosyltransferase localized in the endoplasmic reticulum, significantly increased cell wall chitin content and decreased cell wall β-1,6-glucan content. KRE5 repression induced endoplasmic reticulum stress-related gene expression and MAP kinase pathway activation, including Slt2p and Hog1p phosphorylation, through the cell wall integrity signaling pathway. Moreover, the calcineurin pathway negatively regulated cell wall integrity, but not the reduction of β-1,6-glucan content. These results indicate that KRE5 is required for maintaining both endoplasmic reticulum homeostasis and cell wall integrity, and that the calcineurin pathway acts as a regulator of chitin-glucan balance in the cell wall and as an alternative mediator of endoplasmic reticulum stress in C. glabrata. PMID:27548283

Expert Opinion Editorial Tissue Engineered Blood Vessels as Promising Tools for Testing Drug Toxicity

PubMed Central

Truskey, George A.; Fernandez, Cristina E.

2015-01-01

Drug-induced vascular injury (DIVI) is a serious problem in preclinical studies of vasoactive molecules and for survivors of pediatric cancers. DIVI is often observed in rodents and some larger animals, primarily with drugs affecting vascular tone, but not in humans; however, DIVI observed in animal studies often precludes a drug candidate from continuing along the development pipeline. Thus, there is great interest by the pharmaceutical industry to identify quantifiable human biomarkers of DIVI. Small scale endothelialized tissue-engineered blood vessels using human cells represent a promising approach to screen drug candidates and developed alternatives to cancer therapeutics in vitro. We identify several technical challenges that remain to be addressed, including high throughput systems to screen large numbers of candidates, identification of suitable cell sources, and establishing and maintaining a differentiated state of the vessel wall cells. Adequately addressing these challenges should yield novel platforms to screen drugs and develop new therapeutics to treat cardiovascular disease. PMID:26028128

A poroelastic model describing nutrient transport and cell stresses within a cyclically strained collagen hydrogel.

PubMed

Vaughan, Benjamin L; Galie, Peter A; Stegemann, Jan P; Grotberg, James B

2013-11-05

In the creation of engineered tissue constructs, the successful transport of nutrients and oxygen to the contained cells is a significant challenge. In highly porous scaffolds subject to cyclic strain, the mechanical deformations can induce substantial fluid pressure gradients, which affect the transport of solutes. In this article, we describe a poroelastic model to predict the solid and fluid mechanics of a highly porous hydrogel subject to cyclic strain. The model was validated by matching the predicted penetration of a bead into the hydrogel from the model with experimental observations and provides insight into nutrient transport. Additionally, the model provides estimates of the wall-shear stresses experienced by the cells embedded within the scaffold. These results provide insight into the mechanics of and convective nutrient transport within a cyclically strained hydrogel, which could lead to the improved design of engineered tissues. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Enhanced pathway efficiency of Saccharomyces cerevisiae by introducing thermo-tolerant devices.

PubMed

Liu, Yueqin; Zhang, Genli; Sun, Huan; Sun, Xiangying; Jiang, Nisi; Rasool, Aamir; Lin, Zhanglin; Li, Chun

2014-10-01

In this study, thermo-tolerant devices consisting of heat shock genes from thermophiles were designed and introduced into Saccharomyces cerevisiae for improving its thermo-tolerance. Among ten engineered thermo-tolerant yeasts, T.te-TTE2469, T.te-GroS2 and T.te-IbpA displayed over 25% increased cell density and 1.5-4-fold cell viability compared with the control. Physiological characteristics of thermo-tolerant strains revealed that better cell wall integrity, higher trehalose content and enhanced metabolic energy were preserved by thermo-tolerant devices. Engineered thermo-tolerant strain was used to investigate the impact of thermo-tolerant device on pathway efficiency by introducing β-amyrin synthesis pathway, showed 28.1% increased β-amyrin titer, 28-35°C broadened growth temperature range and 72h shortened fermentation period. The results indicated that implanting heat shock proteins from thermophiles to S. cerevisiae would be an efficient approach to improve its thermo-tolerance. Copyright © 2014 Elsevier Ltd. All rights reserved.

Imaging the Dynamics of Cell Wall Polymer Deposition in the Unicellular Model Plant, Penium margaritaceum.

PubMed

Domozych, David; Lietz, Anna; Patten, Molly; Singer, Emily; Tinaz, Berke; Raimundo, Sandra C

2017-01-01

The unicellular green alga, Penium margaritaceum, represents a novel and valuable model organism for elucidating cell wall dynamics in plants. This organism's cell wall contains several polymers that are highly similar to those found in the primary cell walls of land plants. Penium is easily grown in laboratory culture and is effectively manipulated in various experimental protocols including microplate assays and correlative microscopy. Most importantly, Penium can be live labeled with cell wall-specific antibodies or other probes and returned to culture where specific cell wall developmental events can be monitored. Additionally, live cells can be rapidly cryo-fixed and cell wall surface microarchitecture can be observed with variable pressure scanning electron microscopy. Here, we describe the methodology for maintaining Penium for experimental cell wall enzyme studies.

Cell walls of the dimorphic fungal pathogens Sporothrix schenckii and Sporothrix brasiliensis exhibit bilaminate structures and sloughing of extensive and intact layers

PubMed Central

Walker, Louise A.; Niño-Vega, Gustavo; Mora-Montes, Héctor M.; Neves, Gabriela W. P.; Villalobos-Duno, Hector; Barreto, Laura; Garcia, Karina; Franco, Bernardo; Martínez-Álvarez, José A.; Munro, Carol A.; Gow, Neil A. R.

2018-01-01

Sporotrichosis is a subcutaneous mycosis caused by pathogenic species of the Sporothrix genus. A new emerging species, Sporothrix brasiliensis, is related to cat-transmitted sporotrichosis and has severe clinical manifestations. The cell wall of pathogenic fungi is a unique structure and impacts directly on the host immune response. We reveal and compare the cell wall structures of Sporothrix schenckii and S. brasiliensis using high-pressure freezing electron microscopy to study the cell wall organization of both species. To analyze the components of the cell wall, we also used infrared and 13C and 1H NMR spectroscopy and the sugar composition was determined by quantitative high-performance anion-exchange chromatography. Our ultrastructural data revealed a bi-layered cell wall structure for both species, including an external microfibrillar layer and an inner electron-dense layer. The inner and outer layers of the S. brasiliensis cell wall were thicker than those of S. schenckii, correlating with an increase in the chitin and rhamnose contents. Moreover, the outer microfibrillar layer of the S. brasiliensis cell wall had longer microfibrils interconnecting yeast cells. Distinct from those of other dimorphic fungi, the cell wall of Sporothrix spp. lacked α-glucan component. Interestingly, glycogen α-particles were identified in the cytoplasm close to the cell wall and the plasma membrane. The cell wall structure as well as the presence of glycogen α-particles varied over time during cell culture. The structural differences observed in the cell wall of these Sporothrix species seemed to impact its uptake by monocyte-derived human macrophages. The data presented here show a unique cell wall structure of S. brasiliensis and S. schenckii during the yeast parasitic phase. A new cell wall model for Sporothrix spp. is therefore proposed that suggests that these fungi molt sheets of intact cell wall layers. This observation may have significant effects on localized and disseminated immunopathology. PMID:29522522

DOE Office of Scientific and Technical Information (OSTI.GOV)

Somerville, Chris

Summer Lecture Series 2007: Chris Somerville, Director of the Energy Biosciences Institute and an award-winning plant biochemist with Berkeley Lab's Physical Biosciences Division, is a leading authority on the structure and function of plant cell walls. He discusses an overview of some of the technical challenges associated with the production of cellulosic biofuels, which will require an improved understanding of a diverse range of topics in fields such as agronomy, chemical engineering, microbiology, structural biology, genomics, environmental sciences, and socioeconomics.

Purification and Characterization of the Bacterial Flagellar Basal Body from Salmonella enterica.

PubMed

Aizawa, Shin-Ichi

2017-01-01

The bacterial flagellum is a motility organelle. The flagellum is composed of three main structures: the basal body as a rotary engine embedded in the cellular membranes and cell wall, the long external filament that acts as a propeller, and the hook acting as a universal joint that connects them. I describe protocols for the purification of the filament and hook-basal body from Salmonella enterica serovar Typhimurium.

Optical monitoring system for a turbine engine

DOEpatents

Lemieux, Dennis H; Smed, Jan P; Williams, James P; Jonnalagadda, Vinay

2013-05-14

The monitoring system for a gas turbine engine including a viewing tube assembly having an inner end and an outer end. The inner end is located adjacent to a hot gas flow path within the gas turbine engine and the outer end is located adjacent to an outer casing of the gas turbine engine. An aperture wall is located at the inner end of the viewing tube assembly and an optical element is located within the viewing tube assembly adjacent to the inner end and is spaced from the aperture wall to define a cooling and purge chamber therebetween. An aperture is defined in the aperture wall for passage of light from the hot gas flow path to the optical element. Swirl passages are defined in the viewing tube assembly between the aperture wall and the optical element for passage of cooling air from a location outside the viewing tube assembly into the chamber, wherein swirl passages effect a swirling movement of air in a circumferential direction within the chamber.

Systematic Parameterization of Lignin for the CHARMM Force Field

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vermaas, Joshua; Petridis, Loukas; Beckham, Gregg

Plant cell walls have three primary components, cellulose, hemicellulose, and lignin, the latter of which is a recalcitrant, aromatic heteropolymer that provides structure to plants, water and nutrient transport through plant tissues, and a highly effective defense against pathogens. Overcoming the recalcitrance of lignin is key to effective biomass deconstruction, which would in turn enable the use of biomass as a feedstock for industrial processes. Our understanding of lignin structure in the plant cell wall is hampered by the limitations of the available lignin forcefields, which currently only account for a single linkage between lignins and lack explicit parameterization formore » emerging lignin structures both from natural variants and engineered lignin structures. Since polymerization of lignin occurs via radical intermediates, multiple C-O and C-C linkages have been isolated , and the current force field only represents a small subset of lignin the diverse lignin structures found in plants. In order to take into account the wide range of lignin polymerization chemistries, monomers and dimer combinations of C-, H-, G-, and S-lignins as well as with hydroxycinnamic acid linkages were subjected to extensive quantum mechanical calculations to establish target data from which to build a complete molecular mechanics force field tuned specifically for diverse lignins. This was carried out in a GPU-accelerated global optimization process, whereby all molecules were parameterized simultaneously using the same internal parameter set. By parameterizing lignin specifically, we are able to more accurately represent the interactions and conformations of lignin monomers and dimers relative to a general force field. This new force field will enables computational researchers to study the effects of different linkages on the structure of lignin, as well as construct more accurate plant cell wall models based on observed statistical distributions of lignin that differ between disparate feedstocks, and guide further lignin engineering efforts.« less

Hydraulic Conductivity of Smooth Muscle Cell-Initiated Arterial Cocultures

PubMed Central

Mathura, Rishi A.; Russell-Puleri, Sparkle; Cancel, Limary M.; Tarbell, John M.

2015-01-01

The purpose of the study was to examine the effects of arterial coculture conditions on the transport properties of several in vitro endothelial cell (EC) – smooth muscle cell (SMC) – porous filter constructs in which SMC were grown to confluence first and then EC were inoculated. This order of culturing simulates the environment of a blood vessel wall after endothelial layer damage due to stenting, vascular grafting or other vascular wall insult. For all coculture configurations examined, we observed that hydraulic conductivity (Lp) values were significantly higher than predicted by a resistances-in-series (RIS) model accounting for the Lp of EC and SMC measured separately. The greatest increases were observed when EC were plated directly on top of a confluent SMC layer without an intervening filter, presumably mediated by direct EC – SMC contacts that were observed under confocal microscopy. The results are the opposite of a previous study that showed Lp was significantly reduced compared to an RIS model when EC were grown to confluency first. The physiological, pathophysiological and tissue engineering implications of these results are discussed. PMID:26265460

Hydraulic Conductivity of Smooth Muscle Cell-Initiated Arterial Cocultures.

PubMed

Mathura, Rishi A; Russell-Puleri, Sparkle; Cancel, Limary M; Tarbell, John M

2016-05-01

The purpose of the study was to examine the effects of arterial coculture conditions on the transport properties of several in vitro endothelial cell (EC)-smooth muscle cell (SMC)-porous filter constructs in which SMC were grown to confluence first and then EC were inoculated. This order of culturing simulates the environment of a blood vessel wall after endothelial layer damage due to stenting, vascular grafting or other vascular wall insult. For all coculture configurations examined, we observed that hydraulic conductivity (L(p)) values were significantly higher than predicted by a resistances-in-series (RIS) model accounting for the L(p) of EC and SMC measured separately. The greatest increases were observed when EC were plated directly on top of a confluent SMC layer without an intervening filter, presumably mediated by direct EC-SMC contacts that were observed under confocal microscopy. The results are the opposite of a previous study that showed L(p) was significantly reduced compared to an RIS model when EC were grown to confluency first. The physiological, pathophysiological and tissue engineering implications of these results are discussed.

Three-Dimensional Cell Culture Models for Infectious Disease and Drug Development

NASA Technical Reports Server (NTRS)

Nickerson, Cheryl A.; Honer zu Bentrup, Kerstin; Ott, C. Mark

2005-01-01

Three-dimensional (3-D) cell cultures hold enormous potential to advance our understanding of infectious disease and to effectively translate basic cellular research into clinical applications. Using novel NASA bioreactor technology, the rotating wall vessel (RWV), we have engineered physiologically relevant 3-D human tissue culture models for infectious disease studies. The design of the RWV is based on the understanding that organs and tissues function in a 3-D environment, and that this 3-D architecture is critical for the differentiated form and function of tissues in vivo. The RWV provides large numbers of cells which are amenable to a wide variety of experimental manipulations and provides an easy, reproducible, and cost-effective approach to enhance differentiated features of cell culture models.

Formation of three-dimensional cell/polymer constructs for bone tissue engineering in a spinner flask and a rotating wall vessel bioreactor

NASA Technical Reports Server (NTRS)

Sikavitsas, Vassilios I.; Bancroft, Gregory N.; Mikos, Antonios G.; McIntire, L. V. (Principal Investigator)

2002-01-01

The aim of this study is to investigate the effect of the cell culture conditions of three-dimensional polymer scaffolds seeded with rat marrow stromal cells (MSCs) cultured in different bioreactors concerning the ability of these cells to proliferate, differentiate towards the osteoblastic lineage, and generate mineralized extracellular matrix. MSCs harvested from male Sprague-Dawley rats were culture expanded, seeded on three-dimensional porous 75:25 poly(D,L-lactic-co-glycolic acid) biodegradable scaffolds, and cultured for 21 days under static conditions or in two model bioreactors (a spinner flask and a rotating wall vessel) that enhance mixing of the media and provide better nutrient transport to the seeded cells. The spinner flask culture demonstrated a 60% enhanced proliferation at the end of the first week when compared to static culture. On day 14, all cell/polymer constructs exhibited their maximum alkaline phosphatase activity (AP). Cell/polymer constructs cultured in the spinner flask had 2.4 times higher AP activity than constructs cultured under static conditions on day 14. The total osteocalcin (OC) secretion in the spinner flask culture was 3.5 times higher than the static culture, with a peak OC secretion occurring on day 18. No considerable AP activity and OC secretion were detected in the rotating wall vessel culture throughout the 21-day culture period. The spinner flask culture had the highest calcium content at day 14. On day 21, the calcium deposition in the spinner flask culture was 6.6 times higher than the static cultured constructs and over 30 times higher than the rotating wall vessel culture. Histological sections showed concentration of cells and mineralization at the exterior of the foams at day 21. This phenomenon may arise from the potential existence of nutrient concentration gradients at the interior of the scaffolds. The better mixing provided in the spinner flask, external to the outer surface of the scaffolds, may explain the accelerated proliferation and differentiation of marrow stromal osteoblasts, and the localization of the enhanced mineralization on the external surface of the scaffolds. Copyright 2002 Wiley Periodicals, Inc.

A summary of computational experience at GE Aircraft Engines for complex turbulent flows in gas turbines

NASA Astrophysics Data System (ADS)

Zerkle, Ronald D.; Prakash, Chander

1995-03-01

This viewgraph presentation summarizes some CFD experience at GE Aircraft Engines for flows in the primary gaspath of a gas turbine engine and in turbine blade cooling passages. It is concluded that application of the standard k-epsilon turbulence model with wall functions is not adequate for accurate CFD simulation of aerodynamic performance and heat transfer in the primary gas path of a gas turbine engine. New models are required in the near-wall region which include more physics than wall functions. The two-layer modeling approach appears attractive because of its computational complexity. In addition, improved CFD simulation of film cooling and turbine blade internal cooling passages will require anisotropic turbulence models. New turbulence models must be practical in order to have a significant impact on the engine design process. A coordinated turbulence modeling effort between NASA centers would be beneficial to the gas turbine industry.

A summary of computational experience at GE Aircraft Engines for complex turbulent flows in gas turbines

NASA Technical Reports Server (NTRS)

Zerkle, Ronald D.; Prakash, Chander

1995-01-01

This viewgraph presentation summarizes some CFD experience at GE Aircraft Engines for flows in the primary gaspath of a gas turbine engine and in turbine blade cooling passages. It is concluded that application of the standard k-epsilon turbulence model with wall functions is not adequate for accurate CFD simulation of aerodynamic performance and heat transfer in the primary gas path of a gas turbine engine. New models are required in the near-wall region which include more physics than wall functions. The two-layer modeling approach appears attractive because of its computational complexity. In addition, improved CFD simulation of film cooling and turbine blade internal cooling passages will require anisotropic turbulence models. New turbulence models must be practical in order to have a significant impact on the engine design process. A coordinated turbulence modeling effort between NASA centers would be beneficial to the gas turbine industry.

The receptor-like kinase AtVRLK1 regulates secondary cell wall thickening.

PubMed

Huang, Cheng; Zhang, Rui; Gui, Jinshan; Zhong, Yu; Li, Laigeng

2018-04-20

During the growth and development of land plants, some specialized cells, such as tracheary elements, undergo secondary cell wall thickening. Secondary cell walls contain additional lignin, compared with primary cell walls, thus providing mechanical strength and potentially improving defenses against pathogens. However, the molecular mechanisms that initiate wall thickening are unknown. In this study, we identified an Arabidopsis thaliana leucine-rich repeat receptor-like kinase, encoded by AtVRLK1 (Vascular-Related RLK 1), that is specifically expressed in cells undergoing secondary cell wall thickening. Suppression of AtVRLK1expression resulted in a range of phenotypes that included retarded early elongation of the inflorescence stem, shorter fibers, slower root growth, and shorter flower filaments. In contrast, upregulation of AtVRLK1 led to longer fiber cells, reduced secondary cell wall thickening in fiber and vessel cells, and defects in anther dehiscence. Molecular and cellular analyses showed that downregulation of AtVRLK1 promoted secondary cell wall thickening and upregulation of AtVRLK1 enhanced cell elongation and inhibited secondary cell wall thickening. We propose that AtVRLK1 functions as a signaling component in coordinating cell elongation and cell wall thickening during growth and development. {copyright, serif} 2018 American Society of Plant Biologists. All rights reserved.

Homogenization Models for Carbon Nanotubes

NASA Astrophysics Data System (ADS)

Muc, A.; Jamróz, M.

2004-03-01

Two homogenization models for evaluating Young's modulus of nanocomposites reinforced with single-walled and multi-walled carbon nanotubes are presented. The first model is based on a physical description taking into account the interatomic interaction and nanotube geometry. The elementary cell, here a nanotube with a surrounding resin layer, is treated as a homogeneous body — a material continuum. The second model, similar to a phenomenological engineering one, is obtained by combining the law of mixture with the Cox mechanical model. This model describes the stress distribution along stretched short fibers surrounded by a resin matrix. The similarities between composite materials reinforced with short fibers and nanotubes are elucidated. The results obtained are compared with those for classical microcomposites to demonstrate the advantages and disadvantages of both the composite materials.

Spatio-temporal diversification of the cell wall matrix materials in the developing stomatal complexes of Zea mays.

PubMed

Giannoutsou, E; Apostolakos, P; Galatis, B

2016-11-01

The matrix cell wall materials, in developing Zea mays stomatal complexes are asymmetrically distributed, a phenomenon appearing related to the local cell wall expansion and deformation, the establishment of cell polarity, and determination of the cell division plane. In cells of developing Zea mays stomatal complexes, definite cell wall regions expand determinately and become locally deformed. This differential cell wall behavior is obvious in the guard cell mother cells (GMCs) and the subsidiary cell mother cells (SMCs) that locally protrude towards the adjacent GMCs. The latter, emitting a morphogenetic stimulus, induce polarization/asymmetrical division in SMCs. Examination of immunolabeled specimens revealed that homogalacturonans (HGAs) with a high degree of de-esterification (2F4- and JIM5-HGA epitopes) and arabinogalactan proteins are selectively distributed in the extending and deformed cell wall regions, while their margins are enriched with rhamnogalacturonans (RGAs) containing highly branched arabinans (LM6-RGA epitope). In SMCs, the local cell wall matrix differentiation constitutes the first structural event, indicating the establishment of cell polarity. Moreover, in the premitotic GMCs and SMCs, non-esterified HGAs (2F4-HGA epitope) are preferentially localized in the cell wall areas outlining the cytoplasm where the preprophase band is formed. In these areas, the forthcoming cell plate fuses with the parent cell walls. These data suggest that the described heterogeneity in matrix cell wall materials is probably involved in: (a) local cell wall expansion and deformation, (b) the transduction of the inductive GMC stimulus, and (c) the determination of the division plane in GMCs and SMCs.

Immunogold scanning electron microscopy can reveal the polysaccharide architecture of xylem cell walls

PubMed Central

Sun, Yuliang; Juzenas, Kevin

2017-01-01

Abstract Immunofluorescence microscopy (IFM) and immunogold transmission electron microscopy (TEM) are the two main techniques commonly used to detect polysaccharides in plant cell walls. Both are important in localizing cell wall polysaccharides, but both have major limitations, such as low resolution in IFM and restricted sample size for immunogold TEM. In this study, we have developed a robust technique that combines immunocytochemistry with scanning electron microscopy (SEM) to study cell wall polysaccharide architecture in xylem cells at high resolution over large areas of sample. Using multiple cell wall monoclonal antibodies (mAbs), this immunogold SEM technique reliably localized groups of hemicellulosic and pectic polysaccharides in the cell walls of five different xylem structures (vessel elements, fibers, axial and ray parenchyma cells, and tyloses). This demonstrates its important advantages over the other two methods for studying cell wall polysaccharide composition and distribution in these structures. In addition, it can show the three-dimensional distribution of a polysaccharide group in the vessel lateral wall and the polysaccharide components in the cell wall of developing tyloses. This technique, therefore, should be valuable for understanding the cell wall polysaccharide composition, architecture and functions of diverse cell types. PMID:28398585

Building a plant cell wall at a glance.

PubMed

Lampugnani, Edwin R; Khan, Ghazanfar Abbas; Somssich, Marc; Persson, Staffan

2018-01-29

Plant cells are surrounded by a strong polysaccharide-rich cell wall that aids in determining the overall form, growth and development of the plant body. Indeed, the unique shapes of the 40-odd cell types in plants are determined by their walls, as removal of the cell wall results in spherical protoplasts that are amorphic. Hence, assembly and remodeling of the wall is essential in plant development. Most plant cell walls are composed of a framework of cellulose microfibrils that are cross-linked to each other by heteropolysaccharides. The cell walls are highly dynamic and adapt to the changing requirements of the plant during growth. However, despite the importance of plant cell walls for plant growth and for applications that we use in our daily life such as food, feed and fuel, comparatively little is known about how they are synthesized and modified. In this Cell Science at a Glance article and accompanying poster, we aim to illustrate the underpinning cell biology of the synthesis of wall carbohydrates, and their incorporation into the wall, in the model plant Arabidopsis . © 2018. Published by The Company of Biologists Ltd.

Cell wall evolution and diversity

PubMed Central

Fangel, Jonatan U.; Ulvskov, Peter; Knox, J. P.; Mikkelsen, Maria D.; Harholt, Jesper; Popper, Zoë A.; Willats, William G.T.

2012-01-01

Plant cell walls display a considerable degree of diversity in their compositions and molecular architectures. In some cases the functional significance of a particular cell wall type appears to be easy to discern: secondary cells walls are often reinforced with lignin that provides durability; the thin cell walls of pollen tubes have particular compositions that enable their tip growth; lupin seed cell walls are characteristically thickened with galactan used as a storage polysaccharide. However, more frequently the evolutionary mechanisms and selection pressures that underpin cell wall diversity and evolution are unclear. For diverse green plants (chlorophytes and streptophytes) the rapidly increasing availability of transcriptome and genome data sets, the development of methods for cell wall analyses which require less material for analysis, and expansion of molecular probe sets, are providing new insights into the diversity and occurrence of cell wall polysaccharides and associated biosynthetic genes. Such research is important for refining our understanding of some of the fundamental processes that enabled plants to colonize land and to subsequently radiate so comprehensively. The study of cell wall structural diversity is also an important aspect of the industrial utilization of global polysaccharide bio-resources. PMID:22783271

Reverse Core Engine with Thrust Reverser

NASA Technical Reports Server (NTRS)

Chandler, Jesse M. (Inventor); Suciu, Gabriel L. (Inventor)

2017-01-01

An engine system has a gas generator, a bi-fi wall surrounding at least a portion of the gas generator, a casing surrounding a fan, and the casing having first and second thrust reverser doors which in a deployed position abut each other and the bi-fi wall.

Silencing CHALCONE SYNTHASE in Maize Impedes the Incorporation of Tricin into Lignin and Increases Lignin Content.

PubMed

Eloy, Nubia B; Voorend, Wannes; Lan, Wu; Saleme, Marina de Lyra Soriano; Cesarino, Igor; Vanholme, Ruben; Smith, Rebecca A; Goeminne, Geert; Pallidis, Andreas; Morreel, Kris; Nicomedes, José; Ralph, John; Boerjan, Wout

2017-02-01

Lignin is a phenolic heteropolymer that is deposited in secondary-thickened cell walls, where it provides mechanical strength. A recent structural characterization of cell walls from monocot species showed that the flavone tricin is part of the native lignin polymer, where it is hypothesized to initiate lignin chains. In this study, we investigated the consequences of altered tricin levels on lignin structure and cell wall recalcitrance by phenolic profiling, nuclear magnetic resonance, and saccharification assays of the naturally silenced maize (Zea mays) C2-Idf (inhibitor diffuse) mutant, defective in the CHALCONE SYNTHASE Colorless2 (C2) gene. We show that the C2-Idf mutant produces highly reduced levels of apigenin- and tricin-related flavonoids, resulting in a strongly reduced incorporation of tricin into the lignin polymer. Moreover, the lignin was enriched in β-β and β-5 units, lending support to the contention that tricin acts to initiate lignin chains and that, in the absence of tricin, more monolignol dimerization reactions occur. In addition, the C2-Idf mutation resulted in strikingly higher Klason lignin levels in the leaves. As a consequence, the leaves of C2-Idf mutants had significantly reduced saccharification efficiencies compared with those of control plants. These findings are instructive for lignin engineering strategies to improve biomass processing and biochemical production. © 2017 American Society of Plant Biologists. All Rights Reserved.

Silencing CHALCONE SYNTHASE in Maize Impedes the Incorporation of Tricin into Lignin and Increases Lignin Content

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eloy, Nubia B.; Voorend, Wannes; Lan, Wu

Lignin is a phenolic heteropolymer that is deposited in secondary-thickened cell walls, where it provides mechanical strength. A recent structural characterization of cell walls from monocot species showed that the flavone tricin is part of the native lignin polymer, where it is hypothesized to initiate lignin chains. In this study, we investigated the consequences of altered tricin levels on lignin structure and cell wall recalcitrance by phenolic profiling, nuclear magnetic resonance, and saccharification assays of the naturally silenced maize (Zea mays) C2-Idf (inhibitor diffuse) mutant, defective in the CHALCONE SYNTHASE Colorless2 (C2) gene. We show that the C2-Idf mutant producesmore » highly reduced levels of apigenin- and tricin-related flavonoids, resulting in a strongly reduced incorporation of tricin into the lignin polymer. Moreover, the lignin was enriched in β-β and β-5 units, lending support to the contention that tricin acts to initiate lignin chains and that, in the absence of tricin, more monolignol dimerization reactions occur. In addition, the C2-Idf mutation resulted in strikingly higher Klason lignin levels in the leaves. As a consequence, the leaves of C2-Idf mutants had significantly reduced saccharification efficiencies compared with those of control plants. These findings are instructive for lignin engineering strategies to improve biomass processing and biochemical production.« less

Silencing CHALCONE SYNTHASE in Maize Impedes the Incorporation of Tricin into Lignin and Increases Lignin Content1[OPEN

PubMed Central

2017-01-01

Lignin is a phenolic heteropolymer that is deposited in secondary-thickened cell walls, where it provides mechanical strength. A recent structural characterization of cell walls from monocot species showed that the flavone tricin is part of the native lignin polymer, where it is hypothesized to initiate lignin chains. In this study, we investigated the consequences of altered tricin levels on lignin structure and cell wall recalcitrance by phenolic profiling, nuclear magnetic resonance, and saccharification assays of the naturally silenced maize (Zea mays) C2-Idf (inhibitor diffuse) mutant, defective in the CHALCONE SYNTHASE Colorless2 (C2) gene. We show that the C2-Idf mutant produces highly reduced levels of apigenin- and tricin-related flavonoids, resulting in a strongly reduced incorporation of tricin into the lignin polymer. Moreover, the lignin was enriched in β-β and β-5 units, lending support to the contention that tricin acts to initiate lignin chains and that, in the absence of tricin, more monolignol dimerization reactions occur. In addition, the C2-Idf mutation resulted in strikingly higher Klason lignin levels in the leaves. As a consequence, the leaves of C2-Idf mutants had significantly reduced saccharification efficiencies compared with those of control plants. These findings are instructive for lignin engineering strategies to improve biomass processing and biochemical production. PMID:27940492

Silencing CHALCONE SYNTHASE in Maize Impedes the Incorporation of Tricin into Lignin and Increases Lignin Content

DOE PAGES

Eloy, Nubia B.; Voorend, Wannes; Lan, Wu; ...

2016-12-09

Lignin is a phenolic heteropolymer that is deposited in secondary-thickened cell walls, where it provides mechanical strength. A recent structural characterization of cell walls from monocot species showed that the flavone tricin is part of the native lignin polymer, where it is hypothesized to initiate lignin chains. In this study, we investigated the consequences of altered tricin levels on lignin structure and cell wall recalcitrance by phenolic profiling, nuclear magnetic resonance, and saccharification assays of the naturally silenced maize (Zea mays) C2-Idf (inhibitor diffuse) mutant, defective in the CHALCONE SYNTHASE Colorless2 (C2) gene. We show that the C2-Idf mutant producesmore » highly reduced levels of apigenin- and tricin-related flavonoids, resulting in a strongly reduced incorporation of tricin into the lignin polymer. Moreover, the lignin was enriched in β-β and β-5 units, lending support to the contention that tricin acts to initiate lignin chains and that, in the absence of tricin, more monolignol dimerization reactions occur. In addition, the C2-Idf mutation resulted in strikingly higher Klason lignin levels in the leaves. As a consequence, the leaves of C2-Idf mutants had significantly reduced saccharification efficiencies compared with those of control plants. These findings are instructive for lignin engineering strategies to improve biomass processing and biochemical production.« less

Cell wall targeted in planta iron accumulation enhances biomass conversion and seed iron concentration in Arabidopsis and rice.

PubMed

Yang, Haibing; Wei, Hui; Ma, Guojie; Antunes, Mauricio S; Vogt, Stefan; Cox, Joseph; Zhang, Xiao; Liu, Xiping; Bu, Lintao; Gleber, S Charlotte; Carpita, Nicholas C; Makowski, Lee; Himmel, Michael E; Tucker, Melvin P; McCann, Maureen C; Murphy, Angus S; Peer, Wendy A

2016-10-01

Conversion of nongrain biomass into liquid fuel is a sustainable approach to energy demands as global population increases. Previously, we showed that iron can act as a catalyst to enhance the degradation of lignocellulosic biomass for biofuel production. However, direct addition of iron catalysts to biomass pretreatment is diffusion-limited, would increase the cost and complexity of biorefinery unit operations and may have deleterious environmental impacts. Here, we show a new strategy for in planta accumulation of iron throughout the volume of the cell wall where iron acts as a catalyst in the deconstruction of lignocellulosic biomass. We engineered CBM-IBP fusion polypeptides composed of a carbohydrate-binding module family 11 (CBM11) and an iron-binding peptide (IBP) for secretion into Arabidopsis and rice cell walls. CBM-IBP transformed Arabidopsis and rice plants show significant increases in iron accumulation and biomass conversion compared to respective controls. Further, CBM-IBP rice shows a 35% increase in seed iron concentration and a 40% increase in seed yield in greenhouse experiments. CBM-IBP rice potentially could be used to address iron deficiency, the most common and widespread nutritional disorder according to the World Health Organization. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Engineering E. coli for triglyceride accumulation through native and heterologous metabolic reactions.

PubMed

Rucker, Joanna; Paul, Julie; Pfeifer, Blaine A; Lee, Kyongbum

2013-03-01

Triglycerides, traditionally sourced from plant oils, are heavily used in both industrial and healthcare applications. Commercially significant products produced from triglycerides include biodiesel, lubricants, moisturizers, and oils for cooking and dietary supplements. The need to rely upon plant-based production, however, raises concerns of increasing demand and sustainability. The reliance on crop yields and a strong demand for triglycerides provides motivation to engineer production from a robust microbial platform. In this study, Escherichia coli was engineered to synthesize and accumulate triglycerides. Triglycerides were produced from cell wall phospholipid precursors through engineered expression of two enzymes, phosphatidic acid phosphatase (PAP) and diacylglycerol acyltransferase (DGAT). A liquid chromatography-mass spectrometry (LC-MS) method was developed to analyze the production of triglycerides by the engineered E. coli strains. This proof-of-concept study demonstrated a yield of 1.1 mg/L triglycerides (2 g/L dry cell weight) in lysogeny broth medium containing 5 g/L glucose at 8 h following induction of PAP and DGAT expression. LC-MS results also demonstrated that the intracellular triglyceride composition of E. coli was highly conserved. Triglycerides containing the fatty acid distributions 16:0/16:0/16:1, 16:0/16:0/18:1, and 18:1/16:0/16:1 were found in highest concentrations and represent ∼70 % of triglycerides observed.

Three-dimensional modeling of diesel engine intake flow, combustion and emissions

NASA Technical Reports Server (NTRS)

Reitz, R. D.; Rutland, C. J.

1992-01-01

A three-dimensional computer code (KIVA) is being modified to include state-of-the-art submodels for diesel engine flow and combustion: spray atomization, drop breakup/coalescence, multi-component fuel vaporization, spray/wall interaction, ignition and combustion, wall heat transfer, unburned HC and NOx formation, soot and radiation, and the intake flow process. Improved and/or new submodels which were completed are: wall heat transfer with unsteadiness and compressibility, laminar-turbulent characteristic time combustion with unburned HC and Zeldo'vich NOx, and spray/wall impingement with rebounding and sliding drops. Results to date show that adding the effects of unsteadiness and compressibility improves the accuracy of heat transfer predictions; spray drop rebound can occur from walls at low impingement velocities (e.g., in cold-starting); larger spray drops are formed at the nozzle due to the influence of vaporization on the atomization process; a laminar-and-turbulent characteristic time combustion model has the flexibility to match measured engine combustion data over a wide range of operating conditions; and finally, the characteristic time combustion model can also be extended to allow predictions of ignition. The accuracy of the predictions is being assessed by comparisons with available measurements. Additional supporting experiments are also described briefly. To date, comparisons with measured engine cylinder pressure and heat flux data were made for homogeneous charge, spark-ignited and compression-ignited engines. The model results are in good agreement with the experiments.

Distribution of alginate and cellulose and regulatory role of calcium in the cell wall of the brown alga Ectocarpus siliculosus (Ectocarpales, Phaeophyceae).

PubMed

Terauchi, Makoto; Nagasato, Chikako; Inoue, Akira; Ito, Toshiaki; Motomura, Taizo

2016-08-01

This work investigated a correlation between the three-dimensional architecture and compound-components of the brown algal cell wall. Calcium greatly contributes to the cell wall integrity. Brown algae have a unique cell wall consisting of alginate, cellulose, and sulfated polysaccharides. However, the relationship between the architecture and the composition of the cell wall is poorly understood. Here, we investigated the architecture of the cell wall and the effect of extracellular calcium in the sporophyte and gametophyte of the model brown alga, Ectocarpus siliculosus (Dillwyn) Lyngbye, using transmission electron microscopy, histochemical, and immunohistochemical studies. The lateral cell wall of vegetative cells of the sporophyte thalli had multilayered architecture containing electron-dense and negatively stained fibrils. Electron tomographic analysis showed that the amount of the electron-dense fibrils and the junctions was different between inner and outer layers, and between the perpendicular and tangential directions of the cell wall. By immersing the gametophyte thalli in the low-calcium (one-eighth of the normal concentration) artificial seawater medium, the fibrous layers of the lateral cell wall of vegetative cells became swollen. Destruction of cell wall integrity was also induced by the addition of sorbitol. The results demonstrated that electron-dense fibrils were composed of alginate-calcium fibrous gels, and electron negatively stained fibrils were crystalline cellulose microfibrils. It was concluded that the spatial arrangement of electron-dense fibrils was different between the layers and between the directions of the cell wall, and calcium was necessary for maintaining the fibrous layers in the cell wall. This study provides insights into the design principle of the brown algal cell wall.

Cell Wall Composition and Candidate Biosynthesis Gene Expression During Rice Development.

PubMed

Lin, Fan; Manisseri, Chithra; Fagerström, Alexandra; Peck, Matthew L; Vega-Sánchez, Miguel E; Williams, Brian; Chiniquy, Dawn M; Saha, Prasenjit; Pattathil, Sivakumar; Conlin, Brian; Zhu, Lan; Hahn, Michael G; Willats, William G T; Scheller, Henrik V; Ronald, Pamela C; Bartley, Laura E

2016-10-01

Cell walls of grasses, including cereal crops and biofuel grasses, comprise the majority of plant biomass and intimately influence plant growth, development and physiology. However, the functions of many cell wall synthesis genes, and the relationships among and the functions of cell wall components remain obscure. To better understand the patterns of cell wall accumulation and identify genes that act in grass cell wall biosynthesis, we characterized 30 samples from aerial organs of rice (Oryza sativa cv. Kitaake) at 10 developmental time points, 3-100 d post-germination. Within these samples, we measured 15 cell wall chemical components, enzymatic digestibility and 18 cell wall polysaccharide epitopes/ligands. We also used quantitative reverse transcription-PCR to measure expression of 50 glycosyltransferases, 15 acyltransferases and eight phenylpropanoid genes, many of which had previously been identified as being highly expressed in rice. Most cell wall components vary significantly during development, and correlations among them support current understanding of cell walls. We identified 92 significant correlations between cell wall components and gene expression and establish nine strong hypotheses for genes that synthesize xylans, mixed linkage glucan and pectin components. This work provides an extensive analysis of cell wall composition throughout rice development, identifies genes likely to synthesize grass cell walls, and provides a framework for development of genetically improved grasses for use in lignocellulosic biofuel production and agriculture. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Screening and characterization of plant cell walls using carbohydrate microarrays.

PubMed

Sørensen, Iben; Willats, William G T

2011-01-01

Plant cells are surrounded by cell walls built largely from complex carbohydrates. The primary walls of growing plant cells consist of interdependent networks of three polysaccharide classes: cellulose, cross-linking glycans (also known as hemicelluloses), and pectins. Cellulose microfibrils are tethered together by cross-linking glycans, and this assembly forms the major load-bearing component of primary walls, which is infiltrated with pectic polymers. In the secondary walls of woody tissues, pectins are much reduced and walls are reinforced with the phenolic polymer lignin. Plant cell walls are essential for plant life and also have numerous industrial applications, ranging from wood to nutraceuticals. Enhancing our knowledge of cell wall biology and the effective use of cell wall materials is dependent to a large extent on being able to analyse their fine structures. We have developed a suite of techniques based on microarrays probed with monoclonal antibodies with specificity for cell wall components, and here we present practical protocols for this type of analysis.

Characteristic thickened cell walls of the bracts of the 'eternal flower' Helichrysum bracteatum.

PubMed

Nishikawa, Kuniko; Ito, Hiroaki; Awano, Tatsuya; Hosokawa, Munetaka; Yazawa, Susumu

2008-07-01

Helichrysum bracteatum is called an 'eternal flower' and has large, coloured, scarious bracts. These maintain their aesthetic value without wilting or discoloration for many years. There have been no research studies of cell death or cell morphology of the scarious bract, and hence the aim of this work was to elucidate these characteristics for the bract of H. bracteatum. DAPI (4'6-diamidino-2-phenylindol dihydrochloride) staining and fluorescence microscopy were used for observation of cell nuclei. Light microscopy (LM), transmission electron microscopy (TEM) and polarized light microscopy were used for observation of cells, including cell wall morphology. Cell death occurred at the bract tip during the early stage of flower development. The cell wall was the most prominent characteristic of H. bracteatum bract cells. Characteristic thickened secondary cell walls on the inside of the primary cell walls were observed in both epidermal and inner cells. In addition, the walls of all cells exhibited birefringence. Characteristic thickened secondary cell walls have orientated cellulose microfibrils as well as general secondary cell walls of the tracheary elements. For comparison, these characters were not observed in the petal and bract tissues of Chrysanthemum morifolium. Bracts at anthesis are composed of dead cells. Helichrysum bracteatum bracts have characteristic thickened secondary cell walls that have not been observed in the parenchyma of any other flowers or leaves. The cells of the H. bracteatum bract differ from other tissues with secondary cell walls, suggesting that they may be a new cell type.

Peptidoglycan turnover and recycling in Gram-positive bacteria.

PubMed

Reith, Jan; Mayer, Christoph

2011-10-01

Bacterial cells are protected by an exoskeleton, the stabilizing and shape-maintaining cell wall, consisting of the complex macromolecule peptidoglycan. In view of its function, it could be assumed that the cell wall is a static structure. In truth, however, it is steadily broken down by peptidoglycan-cleaving enzymes during cell growth. In this process, named cell wall turnover, in one generation up to half of the preexisting peptidoglycan of a bacterial cell is released from the wall. This would result in a massive loss of cell material, if turnover products were not be taken up and recovered. Indeed, in the Gram-negative model organism Escherichia coli, peptidoglycan recovery has been recognized as a complex pathway, named cell wall recycling. It involves about a dozen dedicated recycling enzymes that convey cell wall turnover products to peptidoglycan synthesis or energy pathways. Whether Gram-positive bacteria also recover their cell wall is currently questioned. Given the much larger portion of peptidoglycan in the cell wall of Gram-positive bacteria, however, recovery of the wall material would provide an even greater benefit in these organisms compared to Gram-negatives. Consistently, in many Gram-positives, orthologs of recycling enzymes were identified, indicating that the cell wall may also be recycled in these organisms. This mini-review provides a compilation of information about cell wall turnover and recycling in Gram-positive bacteria during cell growth and division, including recent findings relating to muropeptide recovery in Bacillus subtilis and Clostridium acetobutylicum from our group. Furthermore, the impact of cell wall turnover and recycling on biotechnological processes is discussed.

RADIOAUTOGRAPHIC STUDY OF CELL WALL DEPOSITION IN GROWING PLANT CELLS

PubMed Central

Ray, Peter M.

1967-01-01

Segments cut from growing oat coleoptiles and pea stems were fed glucose-3H in presence and absence of the growth hormone indoleacetic acid (IAA). By means of electron microscope radioautography it was demonstrated that new cell wall material is deposited both at the wall surface (apposition) and within the preexisting wall structure (internally). Quantitative profiles for the distribution of incorporation with position through the thickness of the wall were obtained for the thick outer wall of epidermal cells. With both oat coleoptile and pea stem epidermal outer walls, it was found that a larger proportion of the newly synthesized wall material appeared to become incorporated within the wall in the presence of IAA. Extraction experiments on coleoptile tissue showed that activity that had been incorporated into the cell wall interior represented noncellulosic constituents, mainly hemicelluloses, whereas cellulose was deposited largely or entirely by apposition. It seems possible that internal incorporation of hemicelluloses plays a role in the cell wall expansion process that is involved in cell growth. PMID:6064369

Genetic modification of stem cells for transplantation.

PubMed

Phillips, M Ian; Tang, Yao Liang

2008-01-14

Gene modification of cells prior to their transplantation, especially stem cells, enhances their survival and increases their function in cell therapy. Like the Trojan horse, the gene-modified cell has to gain entrance inside the host's walls and survive and deliver its transgene products. Using cellular, molecular and gene manipulation techniques the transplanted cell can be protected in a hostile environment from immune rejection, inflammation, hypoxia and apoptosis. Genetic engineering to modify cells involves constructing modules of functional gene sequences. They can be simple reporter genes or complex cassettes with gene switches, cell specific promoters and multiple transgenes. We discuss methods to deliver and construct gene cassettes with viral and non-viral delivery, siRNA, and conditional Cre/Lox P. We review the current uses of gene-modified stem cells in cardiovascular disease, diabetes, neurological diseases, (including Parkinson's, Alzheimer's and spinal cord injury repair), bone defects, hemophilia, and cancer.

Genetic Modification of Stem Cells for Transplantation

PubMed Central

Phillips, M. Ian; Tang, Yao Liang

2009-01-01

Gene modification of cells for prior to their transplantation, especially stem cells, enhances their survival and increases their function in cell therapy. Like the Trojan horse, the gene modified cell has to gain entrance inside the host’s walls and survive and deliver its transgene products Using cellular, molecular and gene manipulation techniques the transplanted cell can be protected in a hostile environment from immune rejection, inflammation, hypoxia and apoptosis. Genetic engineering to modify cells involves constructing modules of functional gene sequences. They can be simple reporter genes or complex cassettes with gene switches, cell specific promoters and multiple transgenes. We discuss methods to deliver and construct gene cassettes with viral and non viral delivery, siRNA, and conditional Cre/Lox P. We review the current uses of gene modified stem cells in cardiovascular disease, diabetes, neurological diseases,( including Parkinson’s, Alzheimer’s and spinal cord injury repair), bone defects, hemophilia, and cancer. PMID:18031863

Interactions of Condensed Tannins with Saccharomyces cerevisiae Yeast Cells and Cell Walls: Tannin Location by Microscopy.

PubMed

Mekoue Nguela, Julie; Vernhet, Aude; Sieczkowski, Nathalie; Brillouet, Jean-Marc

2015-09-02

Interactions between grape tannins/red wine polyphenols and yeast cells/cell walls was previously studied within the framework of red wine aging and the use of yeast-derived products as an alternative to aging on lees. Results evidenced a quite different behavior between whole cells (biomass grown to elaborate yeast-derived products, inactivated yeast, and yeast inactivated after autolysis) and yeast cell walls (obtained from mechanical disruption of the biomass). Briefly, whole cells exhibited a high capacity to irreversibly adsorb grape and wine tannins, whereas only weak interactions were observed for cell walls. This last point was quite unexpected considering the literature and called into question the real role of cell walls in yeasts' ability to fix tannins. In the present work, tannin location after interactions between grape and wine tannins and yeast cells and cell walls was studied by means of transmission electron microscopy, light epifluorescence, and confocal microscopy. Microscopy observations evidenced that if tannins interact with cell walls, and especially cell wall mannoproteins, they also diffuse freely through the walls of dead cells to interact with their plasma membrane and cytoplasmic components.

The valine and lysine residues in the conserved FxVTxK motif are important for the function of phylogenetically distant plant cellulose synthases.

PubMed

Slabaugh, Erin; Scavuzzo-Duggan, Tess; Chaves, Arielle; Wilson, Liza; Wilson, Carmen; Davis, Jonathan K; Cosgrove, Daniel J; Anderson, Charles T; Roberts, Alison W; Haigler, Candace H

2016-05-01

Cellulose synthases (CESAs) synthesize the β-1,4-glucan chains that coalesce to form cellulose microfibrils in plant cell walls. In addition to a large cytosolic (catalytic) domain, CESAs have eight predicted transmembrane helices (TMHs). However, analogous to the structure of BcsA, a bacterial CESA, predicted TMH5 in CESA may instead be an interfacial helix. This would place the conserved FxVTxK motif in the plant cell cytosol where it could function as a substrate-gating loop as occurs in BcsA. To define the functional importance of the CESA region containing FxVTxK, we tested five parallel mutations in Arabidopsis thaliana CESA1 and Physcomitrella patens CESA5 in complementation assays of the relevant cesa mutants. In both organisms, the substitution of the valine or lysine residues in FxVTxK severely affected CESA function. In Arabidopsis roots, both changes were correlated with lower cellulose anisotropy, as revealed by Pontamine Fast Scarlet. Analysis of hypocotyl inner cell wall layers by atomic force microscopy showed that two altered versions of Atcesa1 could rescue cell wall phenotypes observed in the mutant background line. Overall, the data show that the FxVTxK motif is functionally important in two phylogenetically distant plant CESAs. The results show that Physcomitrella provides an efficient model for assessing the effects of engineered CESA mutations affecting primary cell wall synthesis and that diverse testing systems can lead to nuanced insights into CESA structure-function relationships. Although CESA membrane topology needs to be experimentally determined, the results support the possibility that the FxVTxK region functions similarly in CESA and BcsA. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Analysis of Factors Affecting the Performance of RLV Thrust Cell Liners

NASA Technical Reports Server (NTRS)

Arnold, Steven M. (Technical Monitor); Butler, Daniel T., Jr.; Pinders, Marek-Jerzy

2004-01-01

The reusable launch vehicle (RLV) thrust cell liner, or thrust chamber, is a critical component of the Space Shuttle Main Engine (SSME). It is designed to operate in some of the most severe conditions seen in engineering practice. This requirement, in conjunction with experimentally observed 'dog-house' failure modes characterized by bulging and thinning of the cooling channel wall, provides the motivation to study the factors that influence RLV thrust cell liner performance. Factors or parameters believed to be directly related to the observed characteristic deformation modes leading to failure under in-service loading conditions are identified, and subsequently investigated using the cylindrical version of the higher-order theory for functionally graded materials in conjunction with the Robinson's unified viscoplasticity theory and the power-law creep model for modeling the response of the liner s constituents. Configurations are analyzed in which specific modifications in cooling channel wall thickness or constituent materials are made to determine the influence of these parameters on the deformations resulting in the observed failure modes in the outer walls of the cooling channel. The application of thermal barrier coatings and functional grading are also investigated within this context. Comparison of the higher-order theory results based on the Robinson and power-law creep model predictions has demonstrated that, using the available material parameters, the power-law creep model predicts more precisely the experimentally observed deformation leading to the 'dog-house' failure mode for multiple short cycles, while also providing much improved computational efficiency. However, for a single long cycle, both models predict virtually identical deformations. Increasing the power-law creep model coefficients produces appreciable deformations after just one long cycle that would normally be obtained after multiple cycles, thereby enhancing the efficiency of the analysis. This provides a basis for the development of an accelerated modeling procedure to further characterize dog-house deformation modes in RLV thrust cell liners. Additionally, the results presented herein have demonstrated that the mechanism responsible for deformation leading to 'dog-house' failure modes is driven by pressure, creep/relaxation and geometric effects.

Skeletal muscle derived stem cells microintegrated into a biodegradable elastomer for reconstruction of the abdominal wall.

PubMed

Takanari, Keisuke; Hashizume, Ryotaro; Hong, Yi; Amoroso, Nicholas J; Yoshizumi, Tomo; Gharaibeh, Burhan; Yoshida, Osamu; Nonaka, Kazuhiro; Sato, Hideyoshi; Huard, Johnny; Wagner, William R

2017-01-01

A variety of techniques have been applied to generate tissue engineered constructs, where cells are combined with degradable scaffolds followed by a period of in vitro culture or direct implantation. In the current study, a cellularized scaffold was generated by concurrent deposition of electrospun biodegradable elastomer (poly(ester urethane)urea, PEUU) and electrosprayed culture medium + skeletal muscle-derived stem cells (MDSCs) or electrosprayed culture medium alone as a control. MDSCs were obtained from green fluorescent protein (GFP) transgenic rats. The created scaffolds were implanted into allogenic strain-matched rats to replace a full thickness abdominal wall defect. Both control and MDSC-integrated scaffolds showed extensive cellular infiltration at 4 and 8 wk. The number of blood vessels was higher, the area of residual scaffold was lower, number of multinucleated giant cells was lower and area of connective tissue was lower in MDSC-integrated scaffolds (p < 0.05). GFP + cells co-stained positive for VEGF. Bi-axial mechanical properties of the MDSC-microintegrated constructs better approximated the anisotropic behavior of the native abdominal wall. GFP + cells were observed throughout the scaffold at ∼5% of the cell population at 4 and 8 wk. RNA expression at 4 wk showed higher expression of early myogenic marker Pax7, and b-FGF in the MDSC group. Also, higher expression of myogenin and VEGF were seen in the MDSC group at both 4 and 8 wk time points. The paracrine effect of donor cells on host cells likely contributed to the differences found in vivo between the groups. This approach for the rapid creation of highly-cellularized constructs with soft tissue like mechanics offers an attractive methodology to impart cell-derived bioactivity into scaffolds providing mechanical support during the healing process and might find application in a variety of settings. Copyright © 2016 Elsevier Ltd. All rights reserved.

At the border: the plasma membrane-cell wall continuum.

PubMed

Liu, Zengyu; Persson, Staffan; Sánchez-Rodríguez, Clara

2015-03-01

Plant cells rely on their cell walls for directed growth and environmental adaptation. Synthesis and remodelling of the cell walls are membrane-related processes. During cell growth and exposure to external stimuli, there is a constant exchange of lipids, proteins, and other cell wall components between the cytosol and the plasma membrane/apoplast. This exchange of material and the localization of cell wall proteins at certain spots in the plasma membrane seem to rely on a particular membrane composition. In addition, sensors at the plasma membrane detect changes in the cell wall architecture, and activate cytoplasmic signalling schemes and ultimately cell wall remodelling. The apoplastic polysaccharide matrix is, on the other hand, crucial for preventing proteins diffusing uncontrollably in the membrane. Therefore, the cell wall-plasma membrane link is essential for plant development and responses to external stimuli. This review focuses on the relationship between the cell wall and plasma membrane, and its importance for plant tissue organization. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Gas turbine engine combustor can with trapped vortex cavity

DOEpatents

Burrus, David Louis; Joshi, Narendra Digamber; Haynes, Joel Meier; Feitelberg, Alan S.

2005-10-04

A gas turbine engine combustor can downstream of a pre-mixer has a pre-mixer flowpath therein and circumferentially spaced apart swirling vanes disposed across the pre-mixer flowpath. A primary fuel injector is positioned for injecting fuel into the pre-mixer flowpath. A combustion chamber surrounded by an annular combustor liner disposed in supply flow communication with the pre-mixer. An annular trapped dual vortex cavity located at an upstream end of the combustor liner is defined between an annular aft wall, an annular forward wall, and a circular radially outer wall formed therebetween. A cavity opening at a radially inner end of the cavity is spaced apart from the radially outer wall. Air injection first holes are disposed through the forward wall and air injection second holes are disposed through the aft wall. Fuel injection holes are disposed through at least one of the forward and aft walls.

Plant and algal cell walls: diversity and functionality

PubMed Central

Popper, Zoë A.; Ralet, Marie-Christine; Domozych, David S.

2014-01-01

Background Although plants and many algae (e.g. the Phaeophyceae, brown, and Rhodophyceae, red) are only very distantly related they are united in their possession of carbohydrate-rich cell walls, which are of integral importance being involved in many physiological processes. Furthermore, wall components have applications within food, fuel, pharmaceuticals, fibres (e.g. for textiles and paper) and building materials and have long been an active topic of research. As shown in the 27 papers in this Special Issue, as the major deposit of photosynthetically fixed carbon, and therefore energy investment, cell walls are of undisputed importance to the organisms that possess them, the photosynthetic eukaryotes (plants and algae). The complexities of cell wall components along with their interactions with the biotic and abiotic environment are becoming increasingly revealed. Scope The importance of plant and algal cell walls and their individual components to the function and survival of the organism, and for a number of industrial applications, are illustrated by the breadth of topics covered in this issue, which includes papers concentrating on various plants and algae, developmental stages, organs, cell wall components, and techniques. Although we acknowledge that there are many alternative ways in which the papers could be categorized (and many would fit within several topics), we have organized them as follows: (1) cell wall biosynthesis and remodelling, (2) cell wall diversity, and (3) application of new technologies to cell walls. Finally, we will consider future directions within plant cell wall research. Expansion of the industrial uses of cell walls and potentially novel uses of cell wall components are both avenues likely to direct future research activities. Fundamentally, it is the continued progression from characterization (structure, metabolism, properties and localization) of individual cell wall components through to defining their roles in almost every aspect of plant and algal physiology that will present many of the major challenges in future cell wall research. PMID:25453142

Plant and algal cell walls: diversity and functionality.

PubMed

Popper, Zoë A; Ralet, Marie-Christine; Domozych, David S

2014-10-01

Although plants and many algae (e.g. the Phaeophyceae, brown, and Rhodophyceae, red) are only very distantly related they are united in their possession of carbohydrate-rich cell walls, which are of integral importance being involved in many physiological processes. Furthermore,wall components have applications within food, fuel, pharmaceuticals, fibres (e.g. for textiles and paper) and building materials and have long been an active topic of research. As shown in the 27 papers in this Special Issue, as the major deposit of photosynthetically fixed carbon, and therefore energy investment, cell walls are of undisputed importance to the organisms that possess them, the photosynthetic eukaryotes ( plants and algae). The complexities of cell wall components along with their interactions with the biotic and abiotic environment are becoming increasingly revealed. The importance of plant and algal cell walls and their individual components to the function and survival of the organism, and for a number of industrial applications, are illustrated by the breadth of topics covered in this issue, which includes papers concentrating on various plants and algae, developmental stages, organs, cell wall components, and techniques. Although we acknowledge that there are many alternative ways in which the papers could be categorized (and many would fit within several topics), we have organized them as follows: (1) cell wall biosynthesis and remodelling, (2) cell wall diversity, and (3) application of new technologies to cell walls. Finally, we will consider future directions within plant cell wall research. Expansion of the industrial uses of cell walls and potentially novel uses of cell wall components are both avenues likely to direct future research activities. Fundamentally, it is the continued progression from characterization (structure, metabolism, properties and localization) of individual cell wall components through to defining their roles in almost every aspect of plant and algal physiology that will present many of the major challenges in future cell wall research.

NORTH BASEMENT WALL. IBEAM COLUMNS HAVE BEEN ENCASED IN CONCRETE. ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

NORTH BASEMENT WALL. I-BEAM COLUMNS HAVE BEEN ENCASED IN CONCRETE. STEEL BEAMS LAY ACROSS FIRST FLOOR AWAITING CONCRETE POUR. CAMERA LOOKS SOUTHWEST. INL NEGATIVE NO. 735. Unknown Photographer, 10/6/1950 - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

DBIO Best Thesis Award: Mechanics, Dynamics, and Organization of the Bacterial Cytoskeleton and Cell Wall

NASA Astrophysics Data System (ADS)

Wang, Siyuan

2012-02-01

Bacteria come in a variety of shapes. While the peptidoglycan (PG) cell wall serves as an exoskeleton that defines the static cell shape, the internal bacterial cytoskeleton mediates cell shape by recruiting PG synthesis machinery and thus defining the pattern of cell-wall synthesis. While much is known about the chemistry and biology of the cytoskeleton and cell wall, much of their biophysics, including essential aspects of the functionality, dynamics, and organization, remain unknown. This dissertation aims to elucidate the detailed biophysical mechanisms of cytoskeleton guided wall synthesis. First, I find that the bacterial cytoskeleton MreB contributes nearly as much to the rigidity of an Escherichia coli cell as the cell wall. This conclusion implies that the cytoskeletal polymer MreB applies meaningful force to the cell wall, an idea favored by theoretical modeling of wall growth, and suggests an evolutionary origin of cytoskeleton-governed cell rigidity. Second, I observe that MreB rotates around the long axis of E. coli, and the motion depends on wall synthesis. This is the first discovery of a cell-wall assembly driven molecular motor in bacteria. Third, I prove that both cell-wall synthesis and the PG network have chiral ordering, which is established by the spatial pattern of MreB. This work links the molecular structure of the cytoskeleton and of the cell wall with organismal-scale behavior. Finally, I develop a mathematical model of cytoskeleton-cell membrane interactions, which explains the preferential orientation of different cytoskeleton components in bacteria.

Wall effects in continuous microfluidic magneto-affinity cell separation.

PubMed

Wu, Liqun; Zhang, Yong; Palaniapan, Moorthi; Roy, Partha

2010-05-01

Continuous microfluidic magneto-affinity cell separator combines unique microscale flow phenomenon with advantageous nanobead properties, to isolate cells with high specificity. Owing to the comparable size of the cell-bead complexes and the microchannels, the walls of the microchannel exert a strong influence on the separation of cells by this method. We present a theoretical and experimental study that provides a quantitative description of hydrodynamic wall interactions and wall rolling velocity of cells. A transient convection model describes the transport of cells in two-phase microfluidic flow under the influence of an external magnetic field. Transport of cells along the microchannel walls is also considered via an additional equation. Results show the variation of cell flux in the fluid phases and the wall as a function of a dimensionless parameter arising in the equations. Our results suggest that conditions may be optimized to maximize cell separation while minimizing contact with the wall surfaces. Experimentally measured cell rolling velocities on the wall indicate the presence of other near-wall forces in addition to fluid shear forces. Separation of a human colon carcinoma cell line from a mixture of red blood cells, with folic acid conjugated 1 microm and 200 nm beads, is reported.

Relationship between micro-porosity, water permeability and mechanical behavior in scaffolds for cartilage engineering.

PubMed

Vikingsson, L; Claessens, B; Gómez-Tejedor, J A; Gallego Ferrer, G; Gómez Ribelles, J L

2015-08-01

In tissue engineering the design and optimization of biodegradable polymeric scaffolds with a 3D-structure is an important field. The porous scaffold provide the cells with an adequate biomechanical environment that allows mechanotransduction signals for cell differentiation and the scaffolds also protect the cells from initial compressive loading. The scaffold have interconnected macro-pores that host the cells and newly formed tissue, while the pore walls should be micro-porous to transport nutrients and waste products. Polycaprolactone (PCL) scaffolds with a double micro- and macro-pore architecture have been proposed for cartilage regeneration. This work explores the influence of the micro-porosity of the pore walls on water permeability and scaffold compliance. A Poly(Vinyl Alcohol) with tailored mechanical properties has been used to simulate the growing cartilage tissue inside the scaffold pores. Unconfined and confined compression tests were performed to characterize both the water permeability and the mechanical response of scaffolds with varying size of micro-porosity while volume fraction of the macro-pores remains constant. The stress relaxation tests show that the stress response of the scaffold/hydrogel construct is a synergic effect determined by the performance of the both components. This is interesting since it suggests that the in vivo outcome of the scaffold is not only dependent upon the material architecture but also the growing tissue inside the scaffold׳s pores. On the other hand, confined compression results show that compliance of the scaffold is mainly controlled by the micro-porosity of the scaffold and less by hydrogel density in the scaffold pores. These conclusions bring together valuable information for customizing the optimal scaffold and to predict the in vivo mechanical behavior. Copyright © 2015 Elsevier Ltd. All rights reserved.

SAS-6 engineering reveals interdependence between cartwheel and microtubules in determining centriole architecture.

PubMed

Hilbert, Manuel; Noga, Akira; Frey, Daniel; Hamel, Virginie; Guichard, Paul; Kraatz, Sebastian H W; Pfreundschuh, Moritz; Hosner, Sarah; Flückiger, Isabelle; Jaussi, Rolf; Wieser, Mara M; Thieltges, Katherine M; Deupi, Xavier; Müller, Daniel J; Kammerer, Richard A; Gönczy, Pierre; Hirono, Masafumi; Steinmetz, Michel O

2016-04-01

Centrioles are critical for the formation of centrosomes, cilia and flagella in eukaryotes. They are thought to assemble around a nine-fold symmetric cartwheel structure established by SAS-6 proteins. Here, we have engineered Chlamydomonas reinhardtii SAS-6-based oligomers with symmetries ranging from five- to ten-fold. Expression of a SAS-6 mutant that forms six-fold symmetric cartwheel structures in vitro resulted in cartwheels and centrioles with eight- or nine-fold symmetries in vivo. In combination with Bld10 mutants that weaken cartwheel-microtubule interactions, this SAS-6 mutant produced six- to eight-fold symmetric cartwheels. Concurrently, the microtubule wall maintained eight- and nine-fold symmetries. Expressing SAS-6 with analogous mutations in human cells resulted in nine-fold symmetric centrioles that exhibited impaired length and organization. Together, our data suggest that the self-assembly properties of SAS-6 instruct cartwheel symmetry, and lead us to propose a model in which the cartwheel and the microtubule wall assemble in an interdependent manner to establish the native architecture of centrioles.

Structure, function, and biosynthesis of plant cell walls: proceedings of the seventh annual symposium in botany

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dugger, W.M.; Bartnicki-Garcia, S.

Papers in the following areas were included in these symposium proceedings: (1) cell wall chemistry and biosynthesis; (2) cell wall hydrolysis and associated physiology; (3) cellular events associated with cell wall biosynthesis; and (4) interactions of plant cell walls with pathogens and related responses. Papers have been individually abstracted for the data base. (ACR)

Genetic and environmental modification of the mechanical properties of wood

NASA Astrophysics Data System (ADS)

Sederoff, R.; Allona, I.; Whetten, R.

1996-02-01

Wood is one of the nation's leading raw materials and is used for a wide variety of products, either directly as wood, or as derived materials in pulp and paper. Wood is a biological material and evolved to provide mechanical support and water transport to the early plants that conquered the land. Wood is a tissue that results from the differentiation and programmed cell death of cells that derive from a tissue known as the vascular cambium. The vascular cambium is a thin cylinder of undifferentiated tissue in plant stems and roots that gives rise to several different cell types. Cells that differentiate on the internal side of the cambium form xylem, a tissue composed in major part, of long thin cells that die leaving a network of interconnected cell walls that serve to transport water and to provide mechanical support for the woody plant. The shape and chemical composition of the cells in xylem are well suited for these functions. The structure of cells in xylem determines the mechanical properties of the wood because of the strength derived from the reinforced matrix of the wall. The hydrophobic phenolic surface of the inside of the cell walls is essential to maintain surface tension upon which water transport is based and to resist decay caused by microorganisms. The properties of wood derived from the function of xylem also determine its structural and chemical properties as wood and paper products. Therefore, the physical and chemical properties of wood and paper products also depend on the morphology and composition of the cells from which they are derived. Wood (xylem cell walls) is an anisotropic material, a composite of lignocellulose. It is a matrix of cellulose microfibrils, complexed with hemicelluloses, (carbohydrate polymers which contain sugars other than glucose, both pentoses and hexoses), embedded together in a phenolic matrix of lignin. The high tensile strength of wood in the longitudinal direction, is due to the structure of cellulose and the orientation of the cellulose microfibrils. Lignin provides the embedding matrix that imparts compressive strength and flexibility. The water conducting cells in xylem, the tracheids, are long thin cells, which become the fibers of paper when the lignin is removed from wood during the papermaking process. The length of the tracheids and the thickness of the walls have important effects on the properties of paper that is produced. The past two decades have marked a revolutionary period in biological sciences due to the development of gene splicing techniques. These methods have led to the directed engineering of organisms to develop new industrial products. The technology has been used to produce a wide variety of new pharmaceuticals and transgenic plants and animals. This technology is now also being applied to forest trees.

Transient Simulation of Accumulating Particle Deposition in Pipe Flow

NASA Astrophysics Data System (ADS)

Hewett, James; Sellier, Mathieu

2015-11-01

Colloidal particles that deposit in pipe systems can lead to fouling which is an expensive problem in both the geothermal and oil & gas industries. We investigate the gradual accumulation of deposited colloids in pipe flow using numerical simulations. An Euler-Lagrangian approach is employed for modelling the fluid and particle phases. Particle transport to the pipe wall is modelled with Brownian motion and turbulent diffusion. A two-way coupling exists between the fouled material and the pipe flow; the local mass flux of depositing particles is affected by the surrounding fluid in the near-wall region. This coupling is modelled by changing the cells from fluid to solid as the deposited particles exceed each local cell volume. A similar method has been used to model fouling in engine exhaust systems (Paz et al., Heat Transfer Eng., 34(8-9):674-682, 2013). We compare our deposition velocities and deposition profiles with an experiment on silica scaling in turbulent pipe flow (Kokhanenko et al., 19th AFMC, 2014).

Engineering the cell wall by reducing de-methyl-esterified homogalacturonan improves saccharification of plant tissues for bioconversion.

PubMed

Lionetti, Vincenzo; Francocci, Fedra; Ferrari, Simone; Volpi, Chiara; Bellincampi, Daniela; Galletti, Roberta; D'Ovidio, Renato; De Lorenzo, Giulia; Cervone, Felice

2010-01-12

Plant cell walls represent an abundant, renewable source of biofuel and other useful products. The major bottleneck for the industrial scale-up of their conversion to simple sugars (saccharification), to be subsequently converted by microorganisms into ethanol or other products, is their recalcitrance to enzymatic saccharification. We investigated whether the structure of pectin that embeds the cellulose-hemicellulose network affects the exposure of cellulose to enzymes and consequently the process of saccharification. Reduction of de-methyl-esterified homogalacturonan (HGA) in Arabidopsis plants through the expression of a fungal polygalacturonase (PG) or an inhibitor of pectin methylesterase (PMEI) increased the efficiency of enzymatic saccharification. The improved enzymatic saccharification efficiency observed in transformed plants could also reduce the need for acid pretreatment. Similar results were obtained in PG-expressing tobacco plants and in PMEI-expressing wheat plants, indicating that reduction of de-methyl-esterified HGA may be used in crop species to facilitate the process of biomass saccharification.

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

PubMed Central

Orlean, Peter

2012-01-01

The wall gives a Saccharomyces cerevisiae cell its osmotic integrity; defines cell shape during budding growth, mating, sporulation, and pseudohypha formation; and presents adhesive glycoproteins to other yeast cells. The wall consists of β1,3- and β1,6-glucans, a small amount of chitin, and many different proteins that may bear N- and O-linked glycans and a glycolipid anchor. These components become cross-linked in various ways to form higher-order complexes. Wall composition and degree of cross-linking vary during growth and development and change in response to cell wall stress. This article reviews wall biogenesis in vegetative cells, covering the structure of wall components and how they are cross-linked; the biosynthesis of N- and O-linked glycans, glycosylphosphatidylinositol membrane anchors, β1,3- and β1,6-linked glucans, and chitin; the reactions that cross-link wall components; and the possible functions of enzymatic and nonenzymatic cell wall proteins. PMID:23135325

Retention of Proanthocyanidin in Wine-like Solution Is Conferred by a Dynamic Interaction between Soluble and Insoluble Grape Cell Wall Components.

PubMed

Bindon, Keren A; Li, Sijing; Kassara, Stella; Smith, Paul A

2016-11-09

For better understanding of the factors that impact proanthocyanidin (PA) adsorption by insoluble cell walls or interaction with soluble cell wall-derived components, application of a commercial polygalacturonase enzyme preparation was investigated to modify grape cell wall structure. Soluble and insoluble cell wall material was isolated from the skin and mesocarp components of Vitis vinifera Shiraz grapes. It was observed that significant depolymerization of the insoluble grape cell wall occurred following enzyme application to both grape cell wall fractions, with increased solubilization of rhamnogalacturonan-enriched, low molecular weight polysaccharides. However, in the case of grape mesocarp, the solubilization of protein from cell walls (in buffer) was significant and increased only slightly by the enzyme treatment. Enzyme treatment significantly reduced the adsorption of PA by insoluble cell walls, but this effect was observed only when material solubilized from grape cell walls had been removed. The loss of PA through interaction with the soluble cell wall fraction was observed to be greater for mesocarp than skin cell walls. Subsequent experiments on the soluble mesocarp cell wall fraction confirmed a role for protein in the precipitation of PA. This identified a potential mechanism by which extracted grape PA may be lost from wine during vinification, as a precipitate with solubilized grape mesocarp proteins. Although protein was a minor component in terms of total concentration, losses of PA via precipitation with proteins were in the order of 50% of available PA. PA-induced precipitation could proceed until all protein was removed from solution and may account for the very low levels of residual protein observed in red wines. The results point to a dynamic interaction of grape insoluble and soluble components in modulating PA retention in wine.

The transcription factor Rap1p is required for tolerance to cell-wall perturbing agents and for cell-wall maintenance in Saccharomyces cerevisiae.

PubMed

Azad, Gajendra Kumar; Singh, Vikash; Baranwal, Shivani; Thakare, Mayur Jankiram; Tomar, Raghuvir S

2015-01-02

Yeast repressor activator protein (Rap1p) is involved in genomic stability and transcriptional regulation. We explored the function of Rap1p in yeast physiology using Rap1p truncation mutants. Our results revealed that the N-terminal truncation of Rap1p (Rap1ΔN) leads to hypersensitivity towards elevated temperature and cell-wall perturbing agents. Cell wall analysis showed an increase in the chitin and glucan content in Rap1ΔN cells as compared with wild type cells. Accordingly, mutant cells had a twofold thicker cell wall, as observed by electron microscopy. Furthermore, Rap1ΔN cells had increased levels of phosphorylated Slt2p, a MAP kinase of the cell wall integrity pathway. Mutant cells also had elevated levels of cell wall integrity response transcripts. Taken together, our findings suggest a connection between Rap1p and cell wall homeostasis. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Bacterial cell-wall recycling

PubMed Central

Johnson, Jarrod W.; Fisher, Jed F.; Mobashery, Shahriar

2012-01-01

Many Gram-negative and Gram-positive bacteria recycle a significant proportion of the peptidoglycan components of their cell walls during their growth and septation. In many—and quite possibly all—bacteria, the peptidoglycan fragments are recovered and recycled. While cell-wall recycling is beneficial for the recovery of resources, it also serves as a mechanism to detect cell-wall–targeting antibiotics and to regulate resistance mechanisms. In several Gram-negative pathogens, anhydro-MurNAc-peptide cell-wall fragments regulate AmpC β-lactamase induction. In some Gram-positive organisms, short peptides derived from the cell wall regulate the induction of both β-lactamase and β-lactam-resistant penicillin-binding proteins. The involvement of peptidoglycan recycling with resistance regulation suggests that inhibitors of the enzymes involved in the recycling might synergize with cell-wall-targeted antibiotics. Indeed, such inhibitors improve the potency of β-lactams in vitro against inducible AmpC β-lactamase-producing bacteria. We describe the key steps of cell-wall remodeling and recycling, the regulation of resistance mechanisms by cell-wall recycling, and recent advances toward the discovery of cell-wall recycling inhibitors. PMID:23163477

Characteristic Thickened Cell Walls of the Bracts of the ‘Eternal Flower’ Helichrysum bracteatum

PubMed Central

Nishikawa, Kuniko; Ito, Hiroaki; Awano, Tatsuya; Hosokawa, Munetaka; Yazawa, Susumu

2008-01-01

Background and Aims Helichrysum bracteatum is called an ‘eternal flower’ and has large, coloured, scarious bracts. These maintain their aesthetic value without wilting or discoloration for many years. There have been no research studies of cell death or cell morphology of the scarious bract, and hence the aim of this work was to elucidate these characteristics for the bract of H. bracteatum. Methods DAPI (4'6-diamidino-2-phenylindol dihydrochloride) staining and fluorescence microscopy were used for observation of cell nuclei. Light microscopy (LM), transmission electron microscopy (TEM) and polarized light microscopy were used for observation of cells, including cell wall morphology. Key Results Cell death occurred at the bract tip during the early stage of flower development. The cell wall was the most prominent characteristic of H. bracteatum bract cells. Characteristic thickened secondary cell walls on the inside of the primary cell walls were observed in both epidermal and inner cells. In addition, the walls of all cells exhibited birefringence. Characteristic thickened secondary cell walls have orientated cellulose microfibrils as well as general secondary cell walls of the tracheary elements. For comparison, these characters were not observed in the petal and bract tissues of Chrysanthemum morifolium. Conclusions Bracts at anthesis are composed of dead cells. Helichrysum bracteatum bracts have characteristic thickened secondary cell walls that have not been observed in the parenchyma of any other flowers or leaves. The cells of the H. bracteatum bract differ from other tissues with secondary cell walls, suggesting that they may be a new cell type. PMID:18436550

Mechanical feedback coordinates cell wall expansion and assembly in yeast mating morphogenesis

PubMed Central

2018-01-01

The shaping of individual cells requires a tight coordination of cell mechanics and growth. However, it is unclear how information about the mechanical state of the wall is relayed to the molecular processes building it, thereby enabling the coordination of cell wall expansion and assembly during morphogenesis. Combining theoretical and experimental approaches, we show that a mechanical feedback coordinating cell wall assembly and expansion is essential to sustain mating projection growth in budding yeast (Saccharomyces cerevisiae). Our theoretical results indicate that the mechanical feedback provided by the Cell Wall Integrity pathway, with cell wall stress sensors Wsc1 and Mid2 increasingly activating membrane-localized cell wall synthases Fks1/2 upon faster cell wall expansion, stabilizes mating projection growth without affecting cell shape. Experimental perturbation of the osmotic pressure and cell wall mechanics, as well as compromising the mechanical feedback through genetic deletion of the stress sensors, leads to cellular phenotypes that support the theoretical predictions. Our results indicate that while the existence of mechanical feedback is essential to stabilize mating projection growth, the shape and size of the cell are insensitive to the feedback. PMID:29346368

Wall relaxation and the driving forces for cell expansive growth

NASA Technical Reports Server (NTRS)

Cosgrove, D. J.

1987-01-01

When water uptake by growing cells is prevented, the turgor pressure and the tensile stress in the cell wall are reduced by continued wall loosening. This process, termed in vivo stress relaxation, provides a new way to study the dynamics of wall loosening and to measure the wall yield threshold and the physiological wall extensibility. Stress relaxation experiments indicate that wall stress supplies the mechanical driving force for wall yielding. Cell expansion also requires water absorption. The driving force for water uptake during growth is created by wall relaxation, which lowers the water potential of the expanding cells. New techniques for measuring this driving force show that it is smaller than believed previously; in elongating stems it is only 0.3 to 0.5 bar. This means that the hydraulic resistance of the water transport pathway is small and that rate of cell expansion is controlled primarily by wall loosening and yielding.

Tissue-engineering with muscle fiber fragments improves the strength of a weak abdominal wall in rats.

PubMed

Jangö, Hanna; Gräs, Søren; Christensen, Lise; Lose, Gunnar

2017-02-01

Alternative approaches to reinforce the native tissue in patients with pelvic organ prolapse (POP) are needed to improve surgical outcome. Our aims were to develop a weakened abdominal wall in a rat model to mimic the weakened vaginal wall in women with POP and then evaluate the regenerative potential of a quickly biodegradable synthetic scaffold, methoxypolyethylene glycol polylactic-co-glycolic acid (MPEG-PLGA), seeded with autologous muscle fiber fragments (MFFs) using this model. In an initial pilot study with 15 animals, significant weakening of the abdominal wall and a feasible technique was established by creating a partial defect with removal of one abdominal muscle layer. Subsequently, 18 rats were evenly divided into three groups: (1) unrepaired partial defect; (2) partial defect repaired with MPEG-PLGA; (3) partial defect repaired with MPEG-PLGA and MFFs labeled with PKH26-fluorescence dye. After 8 weeks, we performed histopathological and immunohistochemical testing, fluorescence analysis, and uniaxial biomechanical testing. Both macroscopically and microscopically, the MPEG-PLGA scaffold was fully degraded, with no signs of an inflammatory or foreign-body response. PKH26-positive cells were found in all animals from the group with added MFFs. Analysis of variance (ANOVA) showed a significant difference between groups with respect to load at failure (p = 0.028), and post hoc testing revealed that the group with MPEG-PLGA and MFFs showed a significantly higher strength than the group with MPEG-PLGA alone (p = 0.034). Tissue-engineering with MFFs seeded on a scaffold of biodegradable MPEG-PLGA might be an interesting adjunct to future POP repair.

Two endogenous proteins that induce cell wall extension in plants

NASA Technical Reports Server (NTRS)

McQueen-Mason, S.; Durachko, D. M.; Cosgrove, D. J.

1992-01-01

Plant cell enlargement is regulated by wall relaxation and yielding, which is thought to be catalyzed by elusive "wall-loosening" enzymes. By employing a reconstitution approach, we found that a crude protein extract from the cell walls of growing cucumber seedlings possessed the ability to induce the extension of isolated cell walls. This activity was restricted to the growing region of the stem and could induce the extension of isolated cell walls from various dicot stems and the leaves of amaryllidaceous monocots, but was less effective on grass coleoptile walls. Endogenous and reconstituted wall extension activities showed similar sensitivities to pH, metal ions, thiol reducing agents, proteases, and boiling in methanol or water. Sequential HPLC fractionation of the active wall extract revealed two proteins with molecular masses of 29 and 30 kD associated with the activity. Each protein, by itself, could induce wall extension without detectable hydrolytic breakdown of the wall. These proteins appear to mediate "acid growth" responses of isolated walls and may catalyze plant cell wall extension by a novel biochemical mechanism.

Hydrogels for lung tissue engineering: Biomechanical properties of thin collagen-elastin constructs.

PubMed

Dunphy, Siobhán E; Bratt, Jessica A J; Akram, Khondoker M; Forsyth, Nicholas R; El Haj, Alicia J

2014-10-01

In this study, collagen-elastin constructs were prepared with the aim of producing a material capable of mimicking the mechanical properties of a single alveolar wall. Collagen has been used in a wide range of tissue engineering applications; however, due to its low mechanical properties its use is limited to non load-bearing applications without further manipulation using methods such as cross-linking or mechanical compression. Here, it was hypothesised that the addition of soluble elastin to a collagen hydrogel could improve its mechanical properties. Hydrogels made from collagen only and collagen plus varying amounts elastin were prepared. Young׳s modulus of each membrane was measured using the combination of a non-destructive indentation and a theoretical model previously described. An increase in Young׳s modulus was observed with increasing concentration of elastin. The use of non-destructive indentation allowed for online monitoring of the elastic moduli of cell-seeded constructs over 8 days. The addition of lung fibroblasts into the membrane increased the stiffness of the hydrogels further and cell-seeded collagen hydrogels were found to have a stiffness equal to the theoretical value for a single alveolar wall (≈5kPa). Through provision of some of the native extracellular matrix components of the lung parenchyma these scaffolds may be able to provide an initial building block toward the regeneration of new functional lung tissue. Copyright © 2014 Elsevier Ltd. All rights reserved.

Pectinous cell wall thickenings formation - A common defense strategy of plants to cope with Pb.

PubMed

Krzesłowska, Magdalena; Rabęda, Irena; Basińska, Aneta; Lewandowski, Michał; Mellerowicz, Ewa J; Napieralska, Anna; Samardakiewicz, Sławomir; Woźny, Adam

2016-07-01

Lead, one of the most abundant and hazardous trace metals affecting living organisms, has been commonly detected in plant cell walls including some tolerant plants, mining ecotypes and hyperaccumulators. We have previously shown that in tip growing Funaria sp. protonemata cell wall is remodeled in response to lead by formation of thickenings rich in low-methylesterified pectins (pectin epitope JIM5 - JIM5-P) able to bind metal ions, which accumulate large amounts of Pb. Hence, it leads to the increase of cell wall capacity for Pb compartmentalization. Here we show that diverse plant species belonging to different phyla (Arabidopsis, hybrid aspen, star duckweed), form similar cell wall thickenings in response to Pb. These thickenings are formed in tip growing cells such as the root hairs, and in diffuse growing cells such as meristematic and root cap columella cells of root apices in hybrid aspen and Arabidopsis and in mesophyll cells in star duckweed fronds. Notably, all analyzed cell wall thickenings were abundant in JIM5-P and accumulated high amounts of Pb. In addition, the co-localization of JIM5-P and Pb commonly occurred in these cells. Hence, cell wall thickenings formed the extra compartment for Pb accumulation. In this way plant cells increased cell wall capacity for compartmentalization of this toxic metal, protecting protoplast from its toxicity. As cell wall thickenings occurred in diverse plant species and cell types differing in the type of growth we may conclude that pectinous cell wall thickenings formation is a widespread defense strategy of plants to cope with Pb. Moreover, detection of natural defense strategy, increasing plant cell walls capacity for metal accumulation, reveals a promising direction for enhancing plant efficiency in phytoremediation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Genetic and biochemical characterization of the GH72 family of cell wall transglycosylases in Neurospora crassa.

PubMed

Ao, Jie; Free, Stephen J

2017-04-01

The Neurospora crassa genome encodes five GH72 family transglycosylases, and four of these enzymes (GEL-1, GEL-2, GEL-3 and GEL-5) have been found to be present in the cell wall proteome. We carried out an extensive genetic analysis on the role of these four transglycosylases in cell wall biogenesis and demonstrated that the transglycosylases are required for the formation of a normal cell wall. As suggested by the proteomic analysis, we found that multiple transglycosylases were being expressed in N. crassa cells and that different combinations of the enzymes are required in different cell types. The combination of GEL-1, GEL-2 and GEL-5 is required for the growth of vegetative hyphae, while the GEL-1, GEL-2, GEL-3 combination is needed for the production of aerial hyphae and conidia. Our data demonstrates that the enzymes are redundant with partially overlapping enzymatic activities, which provides the fungus with a robust cell wall biosynthetic system. Characterization of the transglycosylase-deficient mutants demonstrated that the incorporation of cell wall proteins was severely compromised. Interestingly, we found that the transglycosylase-deficient mutant cell walls contained more β-1,3-glucan than the wild type cell wall. Our results demonstrate that the GH72 transglycosylases are not needed for the incorporation of β-1,3-glucan into the cell wall, but they are required for the incorporation of cell wall glycoprotein into the cell wall. Copyright © 2017 Elsevier Inc. All rights reserved.

A Cell Wall Proteome and Targeted Cell Wall Analyses Provide Novel Information on Hemicellulose Metabolism in Flax.

PubMed

Chabi, Malika; Goulas, Estelle; Leclercq, Celine C; de Waele, Isabelle; Rihouey, Christophe; Cenci, Ugo; Day, Arnaud; Blervacq, Anne-Sophie; Neutelings, Godfrey; Duponchel, Ludovic; Lerouge, Patrice; Hausman, Jean-François; Renaut, Jenny; Hawkins, Simon

2017-09-01

Experimentally-generated (nanoLC-MS/MS) proteomic analyses of four different flax organs/tissues (inner-stem, outer-stem, leaves and roots) enriched in proteins from 3 different sub-compartments (soluble-, membrane-, and cell wall-proteins) was combined with publically available data on flax seed and whole-stem proteins to generate a flax protein database containing 2996 nonredundant total proteins. Subsequent multiple analyses (MapMan, CAZy, WallProtDB and expert curation) of this database were then used to identify a flax cell wall proteome consisting of 456 nonredundant proteins localized in the cell wall and/or associated with cell wall biosynthesis, remodeling and other cell wall related processes. Examination of the proteins present in different flax organs/tissues provided a detailed overview of cell wall metabolism and highlighted the importance of hemicellulose and pectin remodeling in stem tissues. Phylogenetic analyses of proteins in the cell wall proteome revealed an important paralogy in the class IIIA xyloglucan endo-transglycosylase/hydrolase (XTH) family associated with xyloglucan endo-hydrolase activity.Immunolocalisation, FT-IR microspectroscopy, and enzymatic fingerprinting indicated that flax fiber primary/S1 cell walls contained xyloglucans with typical substituted side chains as well as glucuronoxylans in much lower quantities. These results suggest a likely central role of xyloglucans and endotransglucosylase/hydrolase activity in flax fiber formation and cell wall remodeling processes. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Developmental and Tissue-Specific Structural Alterations of the Cell-Wall Polysaccharides of Arabidopsis thaliana Roots.

PubMed Central

Freshour, G.; Clay, R. P.; Fuller, M. S.; Albersheim, P.; Darvill, A. G.; Hahn, M. G.

1996-01-01

The plant cell wall is a dynamic structure that plays important roles in growth and development and in the interactions of plants with their environment and other organisms. We have used monoclonal antibodies that recognize different carbohydrate epitopes present in plant cell-wall polysaccharides to locate these epitopes in roots of developing Arabidopsis thaliana seedlings. An epitope in the pectic polysaccharide rhamnogalacturonan I is observed in the walls of epidermal and cortical cells in mature parts of the root. This epitope is inserted into the walls in a developmentally regulated manner. Initially, the epitope is observed in atrichoblasts and later appears in trichoblasts and simultaneously in cortical cells. A terminal [alpha]-fucosyl-containing epitope is present in almost all of the cell walls in the root. An arabinosylated (1->6)-[beta]-galactan epitope is also found in all of the cell walls of the root with the exception of lateral root-cap cell walls. It is striking that these three polysaccharide epitopes are not uniformly distributed (or accessible) within the walls of a given cell, nor are these epitopes distributed equally across the two walls laid down by adjacent cells. Our results further suggest that the biosynthesis and differentiation of primary cell walls in plants are precisely regulated in a temporal, spatial, and developmental manner. PMID:12226270

Atomic force microscopy stiffness tomography on living Arabidopsis thaliana cells reveals the mechanical properties of surface and deep cell-wall layers during growth.

PubMed

Radotić, Ksenija; Roduit, Charles; Simonović, Jasna; Hornitschek, Patricia; Fankhauser, Christian; Mutavdžić, Dragosav; Steinbach, Gabor; Dietler, Giovanni; Kasas, Sandor

2012-08-08

Cell-wall mechanical properties play a key role in the growth and the protection of plants. However, little is known about genuine wall mechanical properties and their growth-related dynamics at subcellular resolution and in living cells. Here, we used atomic force microscopy (AFM) stiffness tomography to explore stiffness distribution in the cell wall of suspension-cultured Arabidopsis thaliana as a model of primary, growing cell wall. For the first time that we know of, this new imaging technique was performed on living single cells of a higher plant, permitting monitoring of the stiffness distribution in cell-wall layers as a function of the depth and its evolution during the different growth phases. The mechanical measurements were correlated with changes in the composition of the cell wall, which were revealed by Fourier-transform infrared (FTIR) spectroscopy. In the beginning and end of cell growth, the average stiffness of the cell wall was low and the wall was mechanically homogenous, whereas in the exponential growth phase, the average wall stiffness increased, with increasing heterogeneity. In this phase, the difference between the superficial and deep wall stiffness was highest. FTIR spectra revealed a relative increase in the polysaccharide/lignin content. Copyright © 2012 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Airfoil for a gas turbine engine

DOEpatents

Liang, George [Palm City, FL

2011-05-24

An airfoil is provided for a turbine of a gas turbine engine. The airfoil comprises: an outer structure comprising a first wall including a leading edge, a trailing edge, a pressure side, and a suction side; an inner structure comprising a second wall spaced from the first wall and at least one intermediate wall; and structure extending between the first and second walls so as to define first and second gaps between the first and second walls. The second wall and the at least one intermediate wall define at least one pressure side supply cavity and at least one suction side supply cavity. The second wall may include at least one first opening near the leading edge of the first wall. The first opening may extend from the at least one pressure side supply cavity to the first gap. The second wall may further comprise at least one second opening near the trailing edge of the outer structure. The second opening may extend from the at least one suction side supply cavity to the second gap. The first wall may comprise at least one first exit opening extending from the first gap through the pressure side of the first wall and at least one second exit opening extending from the second gap through the suction side of the second wall.

Simultaneous co-fermentation of mixed sugars: a promising strategy for producing cellulosic ethanol.

PubMed

Kim, Soo Rin; Ha, Suk-Jin; Wei, Na; Oh, Eun Joong; Jin, Yong-Su

2012-05-01

The lack of microbial strains capable of fermenting all sugars prevalent in plant cell wall hydrolyzates to ethanol is a major challenge. Although naturally existing or engineered microorganisms can ferment mixed sugars (glucose, xylose and galactose) in these hydrolyzates sequentially, the preferential utilization of glucose to non-glucose sugars often results in lower overall yield and productivity of ethanol. Therefore, numerous metabolic engineering approaches have been attempted to construct optimal microorganisms capable of co-fermenting mixed sugars simultaneously. Here, we present recent findings and breakthroughs in engineering yeast for improved ethanol production from mixed sugars. In particular, this review discusses new sugar transporters, various strategies for simultaneous co-fermentation of mixed sugars, and potential applications of co-fermentation for producing fuels and chemicals. Copyright © 2012 Elsevier Ltd. All rights reserved.

Arming Technology in Yeast-Novel Strategy for Whole-cell Biocatalyst and Protein Engineering.

PubMed

Kuroda, Kouichi; Ueda, Mitsuyoshi

2013-09-09

Cell surface display of proteins/peptides, in contrast to the conventional intracellular expression, has many attractive features. This arming technology is especially effective when yeasts are used as a host, because eukaryotic modifications that are often required for functional use can be added to the surface-displayed proteins/peptides. A part of various cell wall or plasma membrane proteins can be genetically fused to the proteins/peptides of interest to be displayed. This technology, leading to the generation of so-called "arming technology", can be employed for basic and applied research purposes. In this article, we describe various strategies for the construction of arming yeasts, and outline the diverse applications of this technology to industrial processes such as biofuel and chemical productions, pollutant removal, and health-related processes, including oral vaccines. In addition, arming technology is suitable for protein engineering and directed evolution through high-throughput screening that is made possible by the feature that proteins/peptides displayed on cell surface can be directly analyzed using intact cells without concentration and purification. Actually, novel proteins/peptides with improved or developed functions have been created, and development of diagnostic/therapeutic antibodies are likely to benefit from this powerful approach.

Improving genetic immobilization of a cellulase on yeast cell surface for bioethanol production using cellulose.

PubMed

Yang, Jinying; Dang, Hongyue; Lu, Jian Ren

2013-04-01

In this study, Saccharomyces cerevisiae was genetically engineered to harbor the capability of utilizing celluloses for bioethanol production by displaying active cellulolytic enzymes on the cell surface. An endo-1,4-β-glucanase gene egX was cloned from Bacillus pumilus C-9 and its expression products, the EGX cellulases, were displayed on the cell surface of S. cerevisiae by fusing egX with aga2 that encodes the binding subunit of the S. cerevisiae cell wall protein α-agglutinin. To achieve high gene copies and stability, multicopy integration was obtained by integrating the fusion aga2-egX gene into the rDNA region of the S. cerevisiae chromosome. To achieve high expression and surface display efficiency, the aga2-egX gene was expressed under the control of a strong promoter. The presence of the enzymatically active cellulase fusion proteins on the S. cerevisiae cell surface was verified by carboxymethyl cellulase activity assay and immunofluorescence microscopy. This work presented a promising strategy to genetically engineer yeasts to perform efficient fermentation of cellulosic materials for bioethanol production. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Fabrication of Cardiac Patch with Decellularized Porcine Myocardial Scaffold and Bone Marrow Mononuclear Cells

PubMed Central

Wang, Bo; Borazjani, Ali; Tahai, Mina; de Jongh Curry, Amy L.; Simionescu, Dan T.; Guan, Jianjun; To, Filip; Elder, Steve H.; Liao, Jun

2010-01-01

Tissue engineered cardiac grafts are a promising therapeutic mode for ventricular wall reconstruction. Recently, it has been found that acellular tissue scaffolds provide natural ultrastructural, mechanical, and compositional cues for recellularization and tissue remodeling. We thus assess the potential of decellularized porcine myocardium as a scaffold for thick cardiac patch tissue engineering. Myocardial sections with 2 mm thickness were decellularized using 0.1% sodium dodecyl sulfate (SDS), and then reseeded with differentiated bone marrow mononuclear cells. We found that thorough decellularization could be achieved after 2.5 weeks treatment. Reseeded cells were found to infiltrate and proliferate in the tissue constructs. Immunohistological staining studies showed that the reseeded cells maintained cardiomyocyte-like phenotype and possible endothelialization was found in locations close to vasculature channels, indicating angiogenesis potential. Both biaxial and uniaxial mechanical testing showed a stiffer mechanical response of the acellular myocardial scaffolds; however, tissue extensibility and tensile modulus were found to recover in the constructs along with the culture time, as expected from increased cellular content. The cardiac patch that we envision for clinical application will benefit from the natural architecture of myocardial extracellular matrix, which has the potential to promote stem cell differentiation, cardiac regeneration, and angiogenesis. PMID:20694977

Shock-Induced Heating In A Rocket Engine

NASA Technical Reports Server (NTRS)

Lagnado, Ronald R.; Raiszadeh, Farhad

1989-01-01

Misalignments give rise to hotspots on walls. Report discusses numerical simulation of flow in and near small, ringlike cavity in wall of Space Shuttle main engine at junction of main combustion chamber and nozzle. Purpose to study effects of misalignments between combustion chamber and nozzle on transfer of heat into surfaces chamber, cavity, and nozzle. Depending on specific misalignment flow encounters forward-or backward-facing step leaving chamber and entering nozzle. Results in serious losses of performance and excessive heating of walls.

Exact solution for flow in a porous pipe with unsteady wall suction and/or injection

NASA Astrophysics Data System (ADS)

Tsangaris, S.; Kondaxakis, D.; Vlachakis, N. W.

2007-10-01

This paper presents an extension of the exact solution of the steady laminar axisymmetric flow in a straight pipe of circular cross section with porous wall, given by R.M. Terrill, to the case of unsteady wall injection and/or suction. The cases of the pulsating parabolic profile and of the developed pulsating flow are investigated as examples. The pulsating flow in porous ducts has many applications in biomedical engineering and in other engineering areas.

A computational approach for inferring the cell wall properties that govern guard cell dynamics.

PubMed

Woolfenden, Hugh C; Bourdais, Gildas; Kopischke, Michaela; Miedes, Eva; Molina, Antonio; Robatzek, Silke; Morris, Richard J

2017-10-01

Guard cells dynamically adjust their shape in order to regulate photosynthetic gas exchange, respiration rates and defend against pathogen entry. Cell shape changes are determined by the interplay of cell wall material properties and turgor pressure. To investigate this relationship between turgor pressure, cell wall properties and cell shape, we focused on kidney-shaped stomata and developed a biomechanical model of a guard cell pair. Treating the cell wall as a composite of the pectin-rich cell wall matrix embedded with cellulose microfibrils, we show that strong, circumferentially oriented fibres are critical for opening. We find that the opening dynamics are dictated by the mechanical stress response of the cell wall matrix, and as the turgor rises, the pectinaceous matrix stiffens. We validate these predictions with stomatal opening experiments in selected Arabidopsis cell wall mutants. Thus, using a computational framework that combines a 3D biomechanical model with parameter optimization, we demonstrate how to exploit subtle shape changes to infer cell wall material properties. Our findings reveal that proper stomatal dynamics are built on two key properties of the cell wall, namely anisotropy in the form of hoop reinforcement and strain stiffening. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd and Society for Experimental Biology.

Virus-induced gene silencing offers a functional genomics platform for studying plant cell wall formation.

PubMed

Zhu, Xiaohong; Pattathil, Sivakumar; Mazumder, Koushik; Brehm, Amanda; Hahn, Michael G; Dinesh-Kumar, S P; Joshi, Chandrashekhar P

2010-09-01

Virus-induced gene silencing (VIGS) is a powerful genetic tool for rapid assessment of plant gene functions in the post-genomic era. Here, we successfully implemented a Tobacco Rattle Virus (TRV)-based VIGS system to study functions of genes involved in either primary or secondary cell wall formation in Nicotiana benthamiana plants. A 3-week post-VIGS time frame is sufficient to observe phenotypic alterations in the anatomical structure of stems and chemical composition of the primary and secondary cell walls. We used cell wall glycan-directed monoclonal antibodies to demonstrate that alteration of cell wall polymer synthesis during the secondary growth phase of VIGS plants has profound effects on the extractability of components from woody stem cell walls. Therefore, TRV-based VIGS together with cell wall component profiling methods provide a high-throughput gene discovery platform for studying plant cell wall formation from a bioenergy perspective.

Plant cell wall signalling and receptor-like kinases.

PubMed

Wolf, Sebastian

2017-02-15

Communication between the extracellular matrix and the cell interior is essential for all organisms as intrinsic and extrinsic cues have to be integrated to co-ordinate development, growth, and behaviour. This applies in particular to plants, the growth and shape of which is governed by deposition and remodelling of the cell wall, a rigid, yet dynamic, extracellular network. It is thus generally assumed that cell wall surveillance pathways exist to monitor the state of the wall and, if needed, elicit compensatory responses such as altered expression of cell wall remodelling and biosynthesis genes. Here, I highlight recent advances in the field of cell wall signalling in plants, with emphasis on the role of plasma membrane receptor-like kinase complexes. In addition, possible roles for cell wall-mediated signalling beyond the maintenance of cell wall integrity are discussed. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

The Specific Nature of Plant Cell Wall Polysaccharides 1

PubMed Central

Nevins, Donald J.; English, Patricia D.; Albersheim, Peter

1967-01-01

Polysaccharide compositions of cell walls were assessed by quantitative analyses of the component sugars. Cell walls were hydrolyzed in 2 n trifluoroacetic acid and the liberated sugars reduced to their respective alditols. The alditols were acetylated and the resulting alditol acetates separated by gas chromatography. Quantitative assay of the alditol acetates was accomplished by electronically integrating the detector output of the gas chromatograph. Myo-inositol, introduced into the sample prior to hydrolysis, served as an internal standard. The cell wall polysaccharide compositions of plant varieties within a given species are essentially identical. However, differences in the sugar composition were observed in cell walls prepared from different species of the same as well as of different genera. The fact that the wall compositions of different varieties of the same species are the same indicates that the biosynthesis of cell wall polysaccharides is genetically regulated. The cell walls of various morphological parts (roots, hypocotyls, first internodes and primary leaves) of bean plants were each found to have a characteristic sugar composition. It was found that the cell wall sugar composition of suspension-cultured sycamore cells could be altered by growing the cells on different carbon sources. This demonstrates that the biosynthesis of cell wall polysaccharides can be manipulated without fatal consequences. PMID:16656594

Remarkable proanthocyanidin adsorption properties of monastrell pomace cell wall material highlight its potential use as an alternative fining agent in red wine production.

PubMed

Bautista-Ortín, Ana Belén; Ruiz-García, Yolanda; Marín, Fátima; Molero, Noelia; Apolinar-Valiente, Rafael; Gómez-Plaza, Encarna

2015-01-21

The existence of interactions between the polysaccharides of vegetal cell walls and proanthocyanins makes this cell wall material an interesting option for its use as a fining agent to reduce the level of proanthocyanins in wines. Pomace wastes from the winery are widely available and a source of cell wall material, and the identification of varieties whose pomace cell walls present high proanthocyanin binding capacity and of processing methods that could enhance their adsorption properties could be of great interest. This study compared the proanthocyanin adsorption properties of pomace cell wall material from three different grape varieties (Monastrell, Cabernet Sauvignon, and Syrah), and the results were compared with those obtained using fresh grape cell walls. Also, the effect of the vinification method has been studied. Analysis of the proanthocyanidins in the solution after reaction with the cell wall material, using phloroglucinolysis and size exclusion chromatography, provided quantitative and qualitative information on the adsorbed and nonadsorbed compounds. A highlight of this study was the observation that Monastrell pomace cell wall material showed a strong affinity for proanthocyanidins, with values similar to that obtained for fresh grapes cell walls, and a preferential binding of high molecular mass proanthocyanidins, so these pomace cell walls could be used in wines to reduce astringency. The use of maceration enzymes during vinification had little effect on the retention capacity of the pomace cell walls obtained from this vinification, although an increase in the retention of low molecular mass proanthocyanidins was observed, and this might have implications for wine sensory properties.

Turbine bucket for use in gas turbine engines and methods for fabricating the same

DOEpatents

Garcia-Crespo, Andres

2014-06-03

A turbine bucket for use with a turbine engine. The turbine bucket includes an airfoil that extends between a root end and a tip end. The airfoil includes an outer wall that defines a cavity that extends from the root end to the tip end. The outer wall includes a first ceramic matrix composite (CMC) substrate that extends a first distance from the root end to the tip end. An inner wall is positioned within the cavity. The inner wall includes a second CMC substrate that extends a second distance from the root end towards the tip end that is different than the first distance.

Apparatus and filtering systems relating to combustors in combustion turbine engines

DOEpatents

Johnson, Thomas Edward [Greer, SC; Zuo, Baifang [Simpsonville, SC; Stevenson, Christian Xavier [Inman, SC

2012-07-24

A combustor for a combustion turbine engine, the combustor that includes: a chamber defined by an outer wall and forming a channel between windows defined through the outer wall toward a forward end of the chamber and at least one fuel injector positioned toward an aft end of the chamber; a screen; and a standoff comprising a raised area on an outer surface of the outer wall near the periphery of the windows; wherein the screen extends over the windows and is supported by the standoff in a raised position in relation to the outer surface of the outer wall and the windows.

Visualizing chemical functionality in plant cell walls

DOE PAGES

Zeng, Yining; Himmel, Michael E.; Ding, Shi-You

2017-11-30

Understanding plant cell wall cross-linking chemistry and polymeric architecture is key to the efficient utilization of biomass in all prospects from rational genetic modification to downstream chemical and biological conversion to produce fuels and value chemicals. In fact, the bulk properties of cell wall recalcitrance are collectively determined by its chemical features over a wide range of length scales from tissue, cellular to polymeric architectures. Microscopic visualization of cell walls from the nanometer to the micrometer scale offers an in situ approach to study their chemical functionality considering its spatial and chemical complexity, particularly the capabilities of characterizing biomass non-destructivelymore » and in real-time during conversion processes. Microscopic characterization has revealed heterogeneity in the distribution of chemical features, which would otherwise be hidden in bulk analysis. Key microscopic features include cell wall type, wall layering, and wall composition - especially cellulose and lignin distributions. Microscopic tools, such as atomic force microscopy, stimulated Raman scattering microscopy, and fluorescence microscopy, have been applied to investigations of cell wall structure and chemistry from the native wall to wall treated by thermal chemical pretreatment and enzymatic hydrolysis. While advancing our current understanding of plant cell wall recalcitrance and deconstruction, microscopic tools with improved spatial resolution will steadily enhance our fundamental understanding of cell wall function.« less

Visualizing chemical functionality in plant cell walls

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zeng, Yining; Himmel, Michael E.; Ding, Shi-You

Understanding plant cell wall cross-linking chemistry and polymeric architecture is key to the efficient utilization of biomass in all prospects from rational genetic modification to downstream chemical and biological conversion to produce fuels and value chemicals. In fact, the bulk properties of cell wall recalcitrance are collectively determined by its chemical features over a wide range of length scales from tissue, cellular to polymeric architectures. Microscopic visualization of cell walls from the nanometer to the micrometer scale offers an in situ approach to study their chemical functionality considering its spatial and chemical complexity, particularly the capabilities of characterizing biomass non-destructivelymore » and in real-time during conversion processes. Microscopic characterization has revealed heterogeneity in the distribution of chemical features, which would otherwise be hidden in bulk analysis. Key microscopic features include cell wall type, wall layering, and wall composition - especially cellulose and lignin distributions. Microscopic tools, such as atomic force microscopy, stimulated Raman scattering microscopy, and fluorescence microscopy, have been applied to investigations of cell wall structure and chemistry from the native wall to wall treated by thermal chemical pretreatment and enzymatic hydrolysis. While advancing our current understanding of plant cell wall recalcitrance and deconstruction, microscopic tools with improved spatial resolution will steadily enhance our fundamental understanding of cell wall function.« less

Visualizing chemical functionality in plant cell walls.

PubMed

Zeng, Yining; Himmel, Michael E; Ding, Shi-You

2017-01-01

Understanding plant cell wall cross-linking chemistry and polymeric architecture is key to the efficient utilization of biomass in all prospects from rational genetic modification to downstream chemical and biological conversion to produce fuels and value chemicals. In fact, the bulk properties of cell wall recalcitrance are collectively determined by its chemical features over a wide range of length scales from tissue, cellular to polymeric architectures. Microscopic visualization of cell walls from the nanometer to the micrometer scale offers an in situ approach to study their chemical functionality considering its spatial and chemical complexity, particularly the capabilities of characterizing biomass non-destructively and in real-time during conversion processes. Microscopic characterization has revealed heterogeneity in the distribution of chemical features, which would otherwise be hidden in bulk analysis. Key microscopic features include cell wall type, wall layering, and wall composition-especially cellulose and lignin distributions. Microscopic tools, such as atomic force microscopy, stimulated Raman scattering microscopy, and fluorescence microscopy, have been applied to investigations of cell wall structure and chemistry from the native wall to wall treated by thermal chemical pretreatment and enzymatic hydrolysis. While advancing our current understanding of plant cell wall recalcitrance and deconstruction, microscopic tools with improved spatial resolution will steadily enhance our fundamental understanding of cell wall function.

The TcEG1 beetle (Tribolium castaneum) cellulase produced in transgenic switchgrass is active at alkaline pH and auto-hydrolyzes biomass for increased cellobiose release

DOE Office of Scientific and Technical Information (OSTI.GOV)

Willis, Jonathan D.; Grant, Joshua N.; Mazarei, Mitra

Genetically engineered biofuel crops, such as switchgrass (Panicum virgatum L.), that produce their own cell wall-digesting cellulase enzymes would reduce costs of cellulosic biofuel production. To date, non-bioenergy plant models have been used in nearly all studies assessing the synthesis and activity of plant-produced fungal and bacterial cellulases. One potential source for cellulolytic enzyme genes is herbivorous insects adapted to digest plant cell walls. Here we examine the potential of transgenic switchgrass-produced TcEG1 cellulase from Tribolium castaneum (red flour beetle). This enzyme, when overproduced in Escherichia coli and Saccharomyces cerevisiae, efficiently digests cellulose at optima of 50 °C and pHmore » 12.0. TcEG1 that was produced in green transgenic switchgrass tissue had a range of endoglucanase activity of 0.16–0.05 units (µM glucose release/min/mg) at 50 °C and pH 12.0. TcEG1 activity from air-dried leaves was unchanged from that from green tissue, but when tissue was dried in a desiccant oven (46 °C), specific enzyme activity decreased by 60%. When transgenic biomass was “dropped-in” into an alkaline buffer (pH 12.0) and allowed to incubate at 50 °C, cellobiose release was increased up to 77% over non-transgenic biomass. Saccharification was increased in one transgenic event by 28%, which had a concurrent decrease in lignin content of 9%. Histological analysis revealed an increase in cell wall thickness with no change to cell area or perimeter. Transgenic plants produced more, albeit narrower, tillers with equivalent dry biomass as the control. This work describes the first study in which an insect cellulase has been produced in transgenic plants; in this case, the dedicated bioenergy crop switchgrass. Switchgrass overexpressing the TcEG1 gene appeared to be morphologically similar to its non-transgenic control and produced equivalent dry biomass. Therefore, we propose TcEG1 transgenics could be bred with other transgenic germplasm (e.g., low-lignin lines) to yield new switchgrass with synergistically reduced recalcitrance to biofuel production. In addition, transgenes for other cell wall degrading enzymes may be stacked with TcEG1 in switchgrass to yield complementary cell wall digestion features and complete auto-hydrolysis.« less

The TcEG1 beetle (Tribolium castaneum) cellulase produced in transgenic switchgrass is active at alkaline pH and auto-hydrolyzes biomass for increased cellobiose release

DOE PAGES

Willis, Jonathan D.; Grant, Joshua N.; Mazarei, Mitra; ...

2017-11-30

Genetically engineered biofuel crops, such as switchgrass (Panicum virgatum L.), that produce their own cell wall-digesting cellulase enzymes would reduce costs of cellulosic biofuel production. To date, non-bioenergy plant models have been used in nearly all studies assessing the synthesis and activity of plant-produced fungal and bacterial cellulases. One potential source for cellulolytic enzyme genes is herbivorous insects adapted to digest plant cell walls. Here we examine the potential of transgenic switchgrass-produced TcEG1 cellulase from Tribolium castaneum (red flour beetle). This enzyme, when overproduced in Escherichia coli and Saccharomyces cerevisiae, efficiently digests cellulose at optima of 50 °C and pHmore » 12.0. TcEG1 that was produced in green transgenic switchgrass tissue had a range of endoglucanase activity of 0.16–0.05 units (µM glucose release/min/mg) at 50 °C and pH 12.0. TcEG1 activity from air-dried leaves was unchanged from that from green tissue, but when tissue was dried in a desiccant oven (46 °C), specific enzyme activity decreased by 60%. When transgenic biomass was “dropped-in” into an alkaline buffer (pH 12.0) and allowed to incubate at 50 °C, cellobiose release was increased up to 77% over non-transgenic biomass. Saccharification was increased in one transgenic event by 28%, which had a concurrent decrease in lignin content of 9%. Histological analysis revealed an increase in cell wall thickness with no change to cell area or perimeter. Transgenic plants produced more, albeit narrower, tillers with equivalent dry biomass as the control. This work describes the first study in which an insect cellulase has been produced in transgenic plants; in this case, the dedicated bioenergy crop switchgrass. Switchgrass overexpressing the TcEG1 gene appeared to be morphologically similar to its non-transgenic control and produced equivalent dry biomass. Therefore, we propose TcEG1 transgenics could be bred with other transgenic germplasm (e.g., low-lignin lines) to yield new switchgrass with synergistically reduced recalcitrance to biofuel production. In addition, transgenes for other cell wall degrading enzymes may be stacked with TcEG1 in switchgrass to yield complementary cell wall digestion features and complete auto-hydrolysis.« less

ETR, TRA642. WALL SECTION DETAILS. METAL SIDING JOINS TO ELECTRICAL ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

ETR, TRA-642. WALL SECTION DETAILS. METAL SIDING JOINS TO ELECTRICAL BUILDING, OFFICE BUILDING, AND ROOF. KAISER ETR-5528-MTR-A-13, 11/1955. INL INDEX NO. 532-0642-00-486-100920, REV. 4. - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

SOUTH WING, MTR661. INTERIOR DETAIL INSIDE LAB ROOM 131. CAMERA ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

SOUTH WING, MTR-661. INTERIOR DETAIL INSIDE LAB ROOM 131. CAMERA FACING NORTHEAST. NOTE CONCRETE BLOCK WALLS. SAFETY SHOWER AND EYE WASHER AT REAR WALL. INL NEGATIVE NO. HD46-7-2. Mike Crane, Photographer, 2/2005. - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

COMPRESSOR BUILDING, TRA626. ELEVATIONS. WINDOWS. WALL SECTIONS. PUMICE BLOCK BUILDING ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

COMPRESSOR BUILDING, TRA-626. ELEVATIONS. WINDOWS. WALL SECTIONS. PUMICE BLOCK BUILDING HOUSED COMPRESSORS FOR AIRCRAFT NUCLEAR PROPULSION EXPERIMENTS. MTR-626-IDO-2S, 3/1952. INL INDEX NO. 531-0626-00-396-110535, REV. 2. - Idaho National Engineering Laboratory, Test Reactor Area, Materials & Engineering Test Reactors, Scoville, Butte County, ID

Formation of wood secondary cell wall may involve two type cellulose synthase complexes in Populus.

PubMed

Xi, Wang; Song, Dongliang; Sun, Jiayan; Shen, Junhui; Li, Laigeng

2017-03-01

Cellulose biosynthesis is mediated by cellulose synthases (CesAs), which constitute into rosette-like cellulose synthase complexe (CSC) on the plasma membrane. Two types of CSCs in Arabidopsis are believed to be involved in cellulose synthesis in the primary cell wall and secondary cell walls, respectively. In this work, we found that the two type CSCs participated cellulose biosynthesis in differentiating xylem cells undergoing secondary cell wall thickening in Populus. During the cell wall thickening process, expression of one type CSC genes increased while expression of the other type CSC genes decreased. Suppression of different type CSC genes both affected the wall-thickening and disrupted the multilaminar structure of the secondary cell walls. When CesA7A was suppressed, crystalline cellulose content was reduced, which, however, showed an increase when CesA3D was suppressed. The CesA suppression also affected cellulose digestibility of the wood cell walls. The results suggest that two type CSCs are involved in coordinating the cellulose biosynthesis in formation of the multilaminar structure in Populus wood secondary cell walls.

Liquid rocket engine nozzles

NASA Technical Reports Server (NTRS)

1976-01-01

The nozzle is a major component of a rocket engine, having a significant influence on the overall engine performance and representing a large fraction of the engine structure. The design of the nozzle consists of solving simultaneously two different problems: the definition of the shape of the wall that forms the expansion surface, and the delineation of the nozzle structure and hydraulic system. This monography addresses both of these problems. The shape of the wall is considered from immediately upstream of the throat to the nozzle exit for both bell and annular (or plug) nozzles. Important aspects of the methods used to generate nozzle wall shapes are covered for maximum-performance shapes and for nozzle contours based on criteria other than performance. The discussion of structure and hydraulics covers problem areas of regeneratively cooled tube-wall nozzles and extensions; it treats also nozzle extensions cooled by turbine exhaust gas, ablation-cooled extensions, and radiation-cooled extensions. The techniques that best enable the designer to develop the nozzle structure with as little difficulty as possible and at the lowest cost consistent with minimum weight and specified performance are described.

Pre-treatment of synthetic elastomeric scaffolds by cardiac fibroblasts improves engineered heart tissue

PubMed Central

Radisic, Milica; Park, Hyoungshin; Martens, Timothy P.; Salazar-Lazaro, Johanna E.; Geng, Wenliang; Wang, Yadong; Langer, Robert; Freed, Lisa E.; Vunjak-Novakovic, Gordana

2009-01-01

Native myocardium consists of several cell types, of which approximately one-third are myocytes and most of the nonmyocytes are fibroblasts. By analogy with monolayer culture in which fibroblasts were removed to prevent overgrowth, early attempts to engineer myocardium utilized cell populations enriched for cardiac myocytes (CMs; ~80–90% of total cells). We hypothesized that the pre-treatment of synthetic elastomeric scaffolds with cardiac fibroblasts (CFs) will enhance the functional assembly of the engineered cardiac constructs by creating an environment supportive of cardiomyocyte attachment and function. Cells isolated from neonatal rat ventricles were prepared to form three distinct populations: rapidly plating cells identified as CFs, slowly plating cells identified as CMs, and unseparated initial population of cells (US). The cell fractions (3 × 106 cells total) were seeded into poly(glycerol sebacate) scaffolds (highly porous discs, 5 mm in diameter × 2-mm thick) using Matrigel™, either separately (CM or CF), concurrently (US), or sequentially (CF pre-treatment followed by CM culture, CF + CM), and cultured in spinner flasks. The CF + CM group had the highest amplitude of contraction and the lowest excitation threshold, superior DNA content, and higher glucose consumption rate. The CF + CM group exhibited compact 100- to 200-μm thick layers of elongated myocytes aligned in parallel over layers of collagen-producing fibroblasts, while US and CM groups exhibited scattered and poorly elongated myocytes. The sequential co-culture of CF and CM on a synthetic elastomer scaffold thus created an environment supportive of cardiomyocyte attachment, differentiation, and contractile function, presumably due to scaffold conditioning by cultured fibroblasts. When implanted over the infarcted myocardium in a nude rat model, cell-free poly(glycerol sebacate) remained at the ventricular wall after 2 weeks of in vivo, and was vascularized. PMID:18041719

Mechanosensation Dynamically Coordinates Polar Growth and Cell Wall Assembly to Promote Cell Survival.

PubMed

Davì, Valeria; Tanimoto, Hirokazu; Ershov, Dmitry; Haupt, Armin; De Belly, Henry; Le Borgne, Rémi; Couturier, Etienne; Boudaoud, Arezki; Minc, Nicolas

2018-04-23

How growing cells cope with size expansion while ensuring mechanical integrity is not known. In walled cells, such as those of microbes and plants, growth and viability are both supported by a thin and rigid encasing cell wall (CW). We deciphered the dynamic mechanisms controlling wall surface assembly during cell growth, using a sub-resolution microscopy approach to monitor CW thickness in live rod-shaped fission yeast cells. We found that polar cell growth yielded wall thinning and that thickness negatively influenced growth. Thickness at growing tips exhibited a fluctuating behavior with thickening phases followed by thinning phases, indicative of a delayed feedback promoting thickness homeostasis. This feedback was mediated by mechanosensing through the CW integrity pathway, which probes strain in the wall to adjust synthase localization and activity to surface growth. Mutants defective in thickness homeostasis lysed by rupturing the wall, demonstrating its pivotal role for walled cell survival. Copyright © 2018 Elsevier Inc. All rights reserved.

The Cell Wall-Associated Proteins in the Dimorphic Pathogenic Species of Paracoccidioides.

PubMed

Puccia, Rosana; Vallejo, Milene C; Longo, Larissa V G

2017-01-01

Paracoccidioides brasiliensis and P. lutzii cause human paracoccidioidomycosis (PCM). They are dimorphic ascomycetes that grow as filaments at mild temperatures up to 28oC and as multibudding pathogenic yeast cells at 37oC. Components of the fungal cell wall have an important role in the interaction with the host because they compose the cell outermost layer. The Paracoccidioides cell wall is composed mainly of polysaccharides, but it also contains proportionally smaller rates of proteins, lipids, and melanin. The polysaccharide cell wall composition and structure of Paracoccidioides yeast cells, filamentous and transition phases were studied in detail in the past. Other cell wall components have been better analyzed in the last decades. The present work gives to the readers a detailed updated view of cell wall-associated proteins. Proteins that have been localized at the cell wall compartment using antibodies are individually addressed. We also make an overview about PCM, the Paracoccidioides cell wall structure, secretion mechanisms, and fungal extracellular vesicles. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Unexpected features of exponentially growing Tobacco Bright Yellow-2 cell suspension culture in relation to excreted extracellular polysaccharides and cell wall composition.

PubMed

Issawi, Mohammad; Muhieddine, Mohammad; Girard, Celine; Sol, Vincent; Riou, Catherine

2017-10-01

This article presents a new insight about TBY-2 cells; from extracellular polysaccharides secretion to cell wall composition during cell suspension culture. In the medium of cells taken 2 days after dilution (end of lag phase), a two unit pH decrease from 5.38 to 3.45 was observed and linked to a high uronic acid (UA) amount secretion (47.8%) while, in 4 and 7 day-old spent media, pH increased and UA amounts decreased 35.6 and 42.3% UA, respectively. To attain deeper knowledge of the putative link between extracellular polysaccharide excretion and cell wall composition, we determined cell wall UA and neutral sugar composition of cells from D2 to D12 cultures. While cell walls from D2 and D3 cells contained a large amount of uronic acid (twice as much as the other analysed cell walls), similar amounts of neutral sugar were detected in cells from lag to end of exponential phase cells suggesting an enriched pectin network in young cultures. Indeed, monosaccharide composition analysis leads to an estimated percentage of pectins of 56% for D3 cell wall against 45% D7 cell walls indicating that the cells at the mid-exponential growth phase re-organized their cell wall linked to a decrease in secreted UA that finally led to a stabilization of the spent medium pH to 5.4. In conclusion, TBY-2 cell suspension from lag to stationary phase showed cell wall remodeling that could be of interest in drug interaction and internalization study.

Investigation of Plant Cell Wall Properties: A Study of Contributions from the Nanoscale to the Macroscale Impacting Cell Wall Recalcitrance

NASA Astrophysics Data System (ADS)

Crowe, Jacob Dillon

Biochemical conversion of lignocellulosic biomass to fuel ethanol is one of a few challenging, yet opportune technologies that can reduce the consumption of petroleum-derived transportation fuels, while providing parallel reductions in greenhouse gas emissions. Biomass recalcitrance, or resistance to deconstruction, is a major technical challenge that limits effective conversion of biomass to fermentable sugars, often requiring a costly thermochemical pretreatment step to improve biomass deconstruction. Biomass recalcitrance is imparted largely by the secondary cell wall, a complex polymeric matrix of cell wall polysaccharides and aromatic heteropolymers, that provides structural stability to cells and enables plant upright growth. Polymers within the cell wall can vary both compositionally and structurally depending upon plant species and anatomical fraction, and have varied responses to thermochemical pretreatments. Cell wall properties impacting recalcitrance are still not well understood, and as a result, the goal of this dissertation is to investigate structural features of the cell wall contributing to recalcitrance (1) in diverse anatomical fractions of a single species, (2) in response to diverse pretreatments, and (3) resulting from genetic modification. In the first study, feedstock cell wall heterogeneity was investigated in anatomical (stem, leaf sheaths, and leaf blades) and internode fractions of switchgrass at varying tissue maturities. Lignin content was observed as the key contributor to recalcitrance in maturing stem tissues only, with non-cellulosic substituted glucuronoarabinoxylans and pectic polysaccharides contributing to cell wall recalcitrance in leaf sheath and leaf blades. Hydroxycinnamate (i.e., saponifiable p-coumarate and ferulate) content along with xylan and pectin extractability decreased with tissue maturity, suggesting lignification is only one component imparting maturity specific cell wall recalcitrance. In the second study, alkaline hydrogen peroxide and liquid hot water pretreatments were shown to alter structural properties impacting nanoscale porosity in corn stover. Delignification by alkaline hydrogen peroxide pretreatment decreased cell wall rigidity, with subsequent cell wall swelling resulting in increased nanoscale porosity and improved enzymatic hydrolysis compared to limited swelling and increased accessible surface areas observed in liquid hot water pretreated biomass. The volume accessible to a 90 A dextran probe within the cell wall was found to be positively correlated to both enzyme binding and glucose hydrolysis yields, indicating cell wall porosity is a key contributor to effective hydrolysis yields. In the third study, the effect of altered xylan content and structure was investigated in irregular xylem (irx) Arabidopsis thaliana mutants to understand the role xylan plays in secondary cell wall development and organization. Higher xylan extractability and lower cellulose crystallinity observed in irx9 and irx15 irx15-L mutants compared to wild type indicated altered xylan integration into the secondary cell wall. Nanoscale cell wall organization observed using multiple microscopy techniques was impacted to some extent in all irx mutants, with disorganized cellulose microfibril layers in sclerenchyma secondary cell walls likely resulting from irregular xylan structure and content. Irregular secondary cell wall microfibril layers showed heterogeneous nanomechanical properties compared to wild type, which translated to mechanical deficiencies observed in stem tensile tests. These results suggest nanoscale defects in cell wall strength can correspond to macroscale phenotypes.

The Acid Growth Theory of auxin-induced cell elongation is alive and well

NASA Technical Reports Server (NTRS)

Rayle, D. L.; Cleland, R. E.

1992-01-01

Plant cells elongate irreversibly only when load-bearing bonds in the walls are cleaved. Auxin causes the elongation of stem and coleoptile cells by promoting wall loosening via cleavage of these bonds. This process may be coupled with the intercalation of new cell wall polymers. Because the primary site of auxin action appears to be the plasma membrane or some intracellular site, and wall loosening is extracellular, there must be communication between the protoplast and the wall. Some "wall-loosening factor" must be exported from auxin-impacted cells, which sets into motion the wall loosening events. About 20 years ago, it was suggested that the wall-loosening factor is hydrogen ions. This idea and subsequent supporting data gave rise to the Acid Growth Theory, which states that when exposed to auxin, susceptible cells excrete protons into the wall (apoplast) at an enhanced rate, resulting in a decrease in apoplastic pH. The lowered wall pH then activates wall-loosening processes, the precise nature of which is unknown. Because exogenous acid causes a transient (1-4 h) increase in growth rate, auxin must also mediate events in addition to wall acidification for growth to continue for an extended period of time. These events may include osmoregulation, cell wall synthesis, and maintenance of the capacity of walls to undergo acid-induced wall loosening. At present, we do not know if these phenomena are tightly coupled to wall acidification or if they are the products of multiple independent signal transduction pathways.

Controlled expression of pectic enzymes in Arabidopsis thaliana enhances biomass conversion without adverse effects on growth.

PubMed

Tomassetti, Susanna; Pontiggia, Daniela; Verrascina, Ilaria; Reca, Ida Barbara; Francocci, Fedra; Salvi, Gianni; Cervone, Felice; Ferrari, Simone

2015-04-01

Lignocellulosic biomass from agriculture wastes is a potential source of biofuel, but its use is currently limited by the recalcitrance of the plant cell wall to enzymatic digestion. Modification of the wall structural components can be a viable strategy to overcome this bottleneck. We have previously shown that the expression of a fungal polygalacturonase (pga2 from Aspergillus niger) in Arabidopsis and tobacco plants reduces the levels of de-esterified homogalacturonan in the cell wall and significantly increases saccharification efficiency. However, plants expressing pga2 show stunted growth and reduced biomass production, likely as a consequence of an extensive loss of pectin integrity during the whole plant life cycle. We report here that the expression in Arabidopsis of another pectic enzyme, the pectate lyase 1 (PL1) of Pectobacterium carotovorum, under the control of a chemically inducible promoter, results, after induction of the transgene, in a saccharification efficiency similar to that of plants expressing pga2. However, lines with high levels of transgene induction show reduced growth even in the absence of the inducer. To overcome the problem of plant fitness, we have generated Arabidopsis plants that express pga2 under the control of the promoter of SAG12, a gene expressed only during senescence. These plants expressed pga2 only at late stages of development, and their growth was comparable to that of WT plants. Notably, leaves and stems of transgenic plants were more easily digested by cellulase, compared to WT plants, only during senescence. Expression of cell wall-degrading enzymes at the end of the plant life cycle may be therefore a useful strategy to engineer crops unimpaired in biomass yield but improved for bioconversion. Copyright © 2014 Elsevier Ltd. All rights reserved.

Cell Wall Ultrastructure of Stem Wood, Roots, and Needles of a Conifer Varies in Response to Moisture Availability

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pattathil, Sivakumar; Ingwers, Miles W.; Victoriano, Olivia L.

The composition, integrity, and architecture of the macromolecular matrix of cell walls, collectively referred to as cell wall ultrastructure, exhibits variation across species and organs and among cell types within organs. Indirect approaches have suggested that modifications to cell wall ultrastructure occur in response to abiotic stress; however, modifications have not been directly observed. Glycome profiling was used to study cell wall ultrastructure by examining variation in composition and extractability of non-cellulosic glycans in cell walls of stem wood, roots, and needles of loblolly pine saplings exposed to high and low soil moisture. Soil moisture influenced physiological processes and themore » overall composition and extractability of cell wall components differed as a function of soil moisture treatments. The strongest response of cell wall ultrastructure to soil moisture was increased extractability of pectic backbone epitopes in the low soil moisture treatment. The higher abundance of these pectic backbone epitopes in the oxalate extract indicate that the loosening of cell wall pectic components could be associated with the release of pectic signals as a stress response. The increased extractability of pectic backbone epitopes in response to low soil moisture availability was more pronounced in stem wood than in roots or needles. Additional responses to low soil moisture availability were observed in lignin associated carbohydrates released in chlorite extracts of stem wood, including an increased abundance of pectic arabinogalactan epitopes. Overall, these results indicate that cell walls of loblolly pine organs undergo changes in their ultrastructural composition and extractability as a response to soil moisture availability and that cell walls of the stem wood are more responsive to low soil moisture availability compared to cell walls of roots and needles. In conclusion, to our knowledge, this is the first direct evidence, delineated by glycomic analyses, that abiotic stress affects cell wall ultrastructure. This study is also unique in that glycome profiling of pine needles has never before been reported.« less

Cell Wall Ultrastructure of Stem Wood, Roots, and Needles of a Conifer Varies in Response to Moisture Availability.

PubMed

Pattathil, Sivakumar; Ingwers, Miles W; Victoriano, Olivia L; Kandemkavil, Sindhu; McGuire, Mary Anne; Teskey, Robert O; Aubrey, Doug P

2016-01-01

The composition, integrity, and architecture of the macromolecular matrix of cell walls, collectively referred to as cell wall ultrastructure, exhibits variation across species and organs and among cell types within organs. Indirect approaches have suggested that modifications to cell wall ultrastructure occur in response to abiotic stress; however, modifications have not been directly observed. Glycome profiling was used to study cell wall ultrastructure by examining variation in composition and extractability of non-cellulosic glycans in cell walls of stem wood, roots, and needles of loblolly pine saplings exposed to high and low soil moisture. Soil moisture influenced physiological processes and the overall composition and extractability of cell wall components differed as a function of soil moisture treatments. The strongest response of cell wall ultrastructure to soil moisture was increased extractability of pectic backbone epitopes in the low soil moisture treatment. The higher abundance of these pectic backbone epitopes in the oxalate extract indicate that the loosening of cell wall pectic components could be associated with the release of pectic signals as a stress response. The increased extractability of pectic backbone epitopes in response to low soil moisture availability was more pronounced in stem wood than in roots or needles. Additional responses to low soil moisture availability were observed in lignin-associated carbohydrates released in chlorite extracts of stem wood, including an increased abundance of pectic arabinogalactan epitopes. Overall, these results indicate that cell walls of loblolly pine organs undergo changes in their ultrastructural composition and extractability as a response to soil moisture availability and that cell walls of the stem wood are more responsive to low soil moisture availability compared to cell walls of roots and needles. To our knowledge, this is the first direct evidence, delineated by glycomic analyses, that abiotic stress affects cell wall ultrastructure. This study is also unique in that glycome profiling of pine needles has never before been reported.

Cell Wall Ultrastructure of Stem Wood, Roots, and Needles of a Conifer Varies in Response to Moisture Availability

DOE PAGES

Pattathil, Sivakumar; Ingwers, Miles W.; Victoriano, Olivia L.; ...

2016-06-24

The composition, integrity, and architecture of the macromolecular matrix of cell walls, collectively referred to as cell wall ultrastructure, exhibits variation across species and organs and among cell types within organs. Indirect approaches have suggested that modifications to cell wall ultrastructure occur in response to abiotic stress; however, modifications have not been directly observed. Glycome profiling was used to study cell wall ultrastructure by examining variation in composition and extractability of non-cellulosic glycans in cell walls of stem wood, roots, and needles of loblolly pine saplings exposed to high and low soil moisture. Soil moisture influenced physiological processes and themore » overall composition and extractability of cell wall components differed as a function of soil moisture treatments. The strongest response of cell wall ultrastructure to soil moisture was increased extractability of pectic backbone epitopes in the low soil moisture treatment. The higher abundance of these pectic backbone epitopes in the oxalate extract indicate that the loosening of cell wall pectic components could be associated with the release of pectic signals as a stress response. The increased extractability of pectic backbone epitopes in response to low soil moisture availability was more pronounced in stem wood than in roots or needles. Additional responses to low soil moisture availability were observed in lignin associated carbohydrates released in chlorite extracts of stem wood, including an increased abundance of pectic arabinogalactan epitopes. Overall, these results indicate that cell walls of loblolly pine organs undergo changes in their ultrastructural composition and extractability as a response to soil moisture availability and that cell walls of the stem wood are more responsive to low soil moisture availability compared to cell walls of roots and needles. In conclusion, to our knowledge, this is the first direct evidence, delineated by glycomic analyses, that abiotic stress affects cell wall ultrastructure. This study is also unique in that glycome profiling of pine needles has never before been reported.« less

Vesicles between plasma membrane and cell wall prior to visible senescence of Iris and Dendrobium flowers.

PubMed

Kamdee, Channatika; Kirasak, Kanjana; Ketsa, Saichol; van Doorn, Wouter G

2015-09-01

Cut Iris flowers (Iris x hollandica, cv. Blue Magic) show visible senescence about two days after full opening. Epidermal cells of the outer tepals collapse due to programmed cell death (PCD). Transmission electron microscopy (TEM) showed irregular swelling of the cell walls, starting prior to cell collapse. Compared to cells in flowers that had just opened, wall thickness increased up to tenfold prior to cell death. Fibrils were visible in the swollen walls. After cell death very little of the cell wall remained. Prior to and during visible wall swelling, vesicles (paramural bodies) were observed between the plasma membrane and the cell walls. The vesicles were also found in groups and were accompanied by amorphous substance. They usually showed a single membrane, and had a variety of diameters and electron densities. Cut Dendrobium hybrid cv. Lucky Duan flowers exhibited visible senescence about 14 days after full flower opening. Paramural bodies were also found in Dendrobium tepal epidermis and mesophyll cells, related to wall swelling and degradation. Although alternative explanations are well possible, it is hypothesized that paramural bodies carry enzymes involved in cell wall breakdown. The literature has not yet reported such bodies in association with senescence/PCD. Copyright © 2015 Elsevier GmbH. All rights reserved.

Changes in cell wall polysaccharide composition, gene transcription and alternative splicing in germinating barley embryos.

PubMed

Zhang, Qisen; Zhang, Xiaoqi; Pettolino, Filomena; Zhou, Gaofeng; Li, Chengdao

2016-02-01

Barley (Hordeum vulgare L.) seed germination initiates many important biological processes such as DNA, membrane and mitochondrial repairs. However, little is known on cell wall modifications in germinating embryos. We have investigated cell wall polysaccharide composition change, gene transcription and alternative splicing events in four barley varieties at 24h and 48 h germination. Cell wall components in germinating barley embryos changed rapidly, with increases in cellulose and (1,3)(1,4)-β-D-glucan (20-100%) within 24h, but decreases in heteroxylan and arabinan (3-50%). There were also significant changes in the levels of type I arabinogalactans and heteromannans. Alternative splicing played very important roles in cell wall modifications. At least 22 cell wall transcripts were detected to undergo either alternative 3' splicing, alternative 5' splicing or intron retention type of alternative splicing. These genes coded enzymes catalyzing synthesis and degradation of cellulose, heteroxylan, (1,3)(1,4)-β-D-glucan and other cell wall polymers. Furthermore, transcriptional regulation also played very important roles in cell wall modifications. Transcript levels of primary wall cellulase synthase, heteroxylan synthesizing and nucleotide sugar inter-conversion genes were very high in germinating embryos. At least 50 cell wall genes changed transcript levels significantly. Expression patterns of many cell wall genes coincided with changes in polysaccharide composition. Our data showed that cell wall polysaccharide metabolism was very active in germinating barley embryos, which was regulated at both transcriptional and post-transcriptional levels. Copyright © 2015 Elsevier GmbH. All rights reserved.

Modelling cell wall growth using a fibre-reinforced hyperelastic-viscoplastic constitutive law

NASA Astrophysics Data System (ADS)

Huang, R.; Becker, A. A.; Jones, I. A.

2012-04-01

A fibre-reinforced hyperelastic-viscoplastic model using a finite strain Finite Element (FE) analysis is presented to study the expansive growth of cell walls. Based on the connections between biological concepts and plasticity theory, e.g. wall-loosening and plastic yield, wall-stiffening and plastic hardening, the modelling of cell wall growth is established within a framework of anisotropic viscoplasticity aiming to represent the corresponding biology-controlled behaviour of a cell wall. In order to model in vivo growth, special attention is paid to the differences between a living cell and an isolated wall. The proposed hyperelastic-viscoplastic theory provides a unique framework to clarify the interplay between cellulose microfibrils and cell wall matrix and how this interplay regulates sustainable growth in a particular direction while maintaining the mechanical strength of the cell walls by new material deposition. Moreover, the effect of temperature is taken into account. A numerical scheme is suggested and FE case studies are presented and compared with experimental data.

Antisense expression of the fasciclin-like arabinogalactan protein FLA6 gene in Populus inhibits expression of its homologous genes and alters stem biomechanics and cell wall composition in transgenic trees.

PubMed

Wang, Haihai; Jiang, Chunmei; Wang, Cuiting; Yang, Yang; Yang, Lei; Gao, Xiaoyan; Zhang, Hongxia

2015-03-01

Fasciclin-like arabinogalactan proteins (FLAs) play important roles in the growth and development of roots, stems, and seeds in Arabidopsis. However, their biological functions in woody plants are largely unknown. In this work, we investigated the possible function of PtFLA6 in poplar. Quantitative real-time PCR, PtFLA6-yellow fluorescent protein (YFP) fusion protein subcellular localization, Western blotting, and immunohistochemical analyses demonstrated that the PtFLA6 gene was expressed specifically in the xylem of mature stem, and PtFLA6 protein was distributed ubiquitous in plant cells and accumulated predominantly in stem xylem fibres. Antisense expression of PtFLA6 in the aspen hybrid clone Poplar davidiana×Poplar bolleana reduced the transcripts of PtFLA6 and its homologous genes. Transgenic plants that showed a significant reduction in the transcripts of PtFLAs accumulated fewer PtFLA6 and arabinogalactan proteins than did the non-transgenic plants, leading to reduced stem flexural strength and stiffness. Further studies revealed that the altered stem biomechanics of transgenic plants could be attributed to the decreased cellulose and lignin composition in the xylem. In addition expression of some xylem-specific genes involved in cell wall biosynthesis was downregulated in these transgenic plants. All these results suggest that engineering the expression of PtFLA6 and its homologues could modulate stem mechanical properties by affecting cell wall composition in trees. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Promoting acid resistance and nisin yield of Lactococcus lactis F44 by genetically increasing D-Asp amidation level inside cell wall.

PubMed

Hao, Panlong; Liang, Dongmei; Cao, Lijie; Qiao, Bin; Wu, Hao; Caiyin, Qinggele; Zhu, Hongji; Qiao, Jianjun

2017-08-01

Nisin fermentation by Lactococcus lactis requires a low pH to maintain a relatively higher nisin activity. However, the acidic environment will result in cell arrest, and eventually decrease the relative nisin production. Hence, constructing an acid-resistant L. lactis is crucial for nisin harvest in acidic nisin fermentation. In this paper, the first discovery of the relationship between D-Asp amidation-associated gene (asnH) and acid resistance was reported. Overexpression of asnH in L. lactis F44 (F44A) resulted in a sevenfold increase in survival capacity during acid shift (pH 3) and enhanced nisin desorption capacity compared to F44 (wild type), which subsequently contributed to higher nisin production, reaching 5346 IU/mL, 57.0% more than that of F44 in the fed-batch fermentation. Furthermore, the engineered F44A showed a moderate increase in D-Asp amidation level (from 82 to 92%) compared to F44. The concomitant decrease of the negative charge inside the cell wall was detected by a newly developed method based on the nisin adsorption amount onto cell surface. Meanwhile, peptidoglycan cross-linkage increased from 36.8% (F44) to 41.9% (F44A), and intracellular pH can be better maintained by blocking extracellular H + due to the maintenance of peptidoglycan integrity, which probably resulted from the action of inhibiting hydrolases activity. The inference was further supported by the acmC-overexpression strain F44C, which was characterized by uncontrolled peptidoglycan hydrolase activity. Our results provided a novel strategy for enhancing nisin yield through cell wall remodeling, which contributed to both continuous nisin synthesis and less nisin adsorption in acidic fermentation (dual enhancement).

(Hydroxyproline-rich glycoproteins of the plant cell wall)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Varner, J.E.

1990-01-01

We are studying the chemistry and architecture of plant cells walls, the extracellular matrices that taken together shape the plant and provide mechanical support for the plant. Cell walls are dynamic structures that regulate, or are the site of, many physiological processes, in addition to being the cells' first line of defense against invading pathogens. In the past year we have examined the role of the cell wall enzyme ascorbic acid oxidase as related to the structure of the wall and its possible interactions with hydroxyproline-rich glycoproteins of the wall.

A Structurally Specialized Uniform Wall Layer is Essential for Constructing Wall Ingrowth Papillae in Transfer Cells

PubMed Central

Xia, Xue; Zhang, Hui-Ming; Offler, Christina E.; Patrick, John W.

2017-01-01

Transfer cells are characterized by wall labyrinths with either a flange or reticulate architecture. A literature survey established that reticulate wall ingrowth papillae ubiquitously arise from a modified component of their wall labyrinth, termed the uniform wall layer; a structure absent from flange transfer cells. This finding sparked an investigation of the deposition characteristics and role of the uniform wall layer using a Vicia faba cotyledon culture system. On transfer of cotyledons to culture, their adaxial epidermal cells spontaneously trans-differentiate to a reticulate architecture comparable to their abaxial epidermal transfer cell counterparts formed in planta. Uniform wall layer construction commenced once adaxial epidermal cell expansion had ceased to overlay the original outer periclinal wall on its inner surface. In contrast to the dense ring-like lattice of cellulose microfibrils in the original primary wall, the uniform wall layer was characterized by a sparsely dispersed array of linear cellulose microfibrils. A re-modeled cortical microtubule array exerted no influence on uniform wall layer formation or on its cellulose microfibril organization. Surprisingly, formation of the uniform wall layer was not dependent upon depositing a cellulose scaffold. In contrast, uniform wall cellulose microfibrils were essential precursors for constructing wall ingrowth papillae. On converging to form wall ingrowth papillae, the cellulose microfibril diameters increased 3-fold. This event correlated with up-regulated differential, and transfer-cell specific, expression of VfCesA3B while transcript levels of other cellulose biosynthetic-related genes linked with primary wall construction were substantially down-regulated. PMID:29259611

A Structurally Specialized Uniform Wall Layer is Essential for Constructing Wall Ingrowth Papillae in Transfer Cells.

PubMed

Xia, Xue; Zhang, Hui-Ming; Offler, Christina E; Patrick, John W

2017-01-01

Transfer cells are characterized by wall labyrinths with either a flange or reticulate architecture. A literature survey established that reticulate wall ingrowth papillae ubiquitously arise from a modified component of their wall labyrinth, termed the uniform wall layer; a structure absent from flange transfer cells. This finding sparked an investigation of the deposition characteristics and role of the uniform wall layer using a Vicia faba cotyledon culture system. On transfer of cotyledons to culture, their adaxial epidermal cells spontaneously trans -differentiate to a reticulate architecture comparable to their abaxial epidermal transfer cell counterparts formed in planta . Uniform wall layer construction commenced once adaxial epidermal cell expansion had ceased to overlay the original outer periclinal wall on its inner surface. In contrast to the dense ring-like lattice of cellulose microfibrils in the original primary wall, the uniform wall layer was characterized by a sparsely dispersed array of linear cellulose microfibrils. A re-modeled cortical microtubule array exerted no influence on uniform wall layer formation or on its cellulose microfibril organization. Surprisingly, formation of the uniform wall layer was not dependent upon depositing a cellulose scaffold. In contrast, uniform wall cellulose microfibrils were essential precursors for constructing wall ingrowth papillae. On converging to form wall ingrowth papillae, the cellulose microfibril diameters increased 3-fold. This event correlated with up-regulated differential, and transfer-cell specific, expression of VfCesA3B while transcript levels of other cellulose biosynthetic-related genes linked with primary wall construction were substantially down-regulated.

Isolation of a novel cell wall architecture mutant of rice with defective Arabidopsis COBL4 ortholog BC1 required for regulated deposition of secondary cell wall components.

PubMed

Sato, Kanna; Suzuki, Ryu; Nishikubo, Nobuyuki; Takenouchi, Sachi; Ito, Sachiko; Nakano, Yoshimi; Nakaba, Satoshi; Sano, Yuzou; Funada, Ryo; Kajita, Shinya; Kitano, Hidemi; Katayama, Yoshihiro

2010-06-01

The plant secondary cell wall is a highly ordered structure composed of various polysaccharides, phenolic components and proteins. Its coordinated regulation of a number of complex metabolic pathways and assembly has not been resolved. To understand the molecular mechanisms that regulate secondary cell wall synthesis, we isolated a novel rice mutant, cell wall architecture1 (cwa1), that exhibits an irregular thickening pattern in the secondary cell wall of sclerenchyma, as well as culm brittleness and reduced cellulose content in mature internodes. Light and transmission electron microscopy revealed that the cwa1 mutant plant has regions of local aggregation in the secondary cell walls of the cortical fibers in its internodes, showing uneven thickness. Ultraviolet microscopic observation indicated that localization of cell wall phenolic components was perturbed and that these components abundantly deposited at the aggregated cell wall regions in sclerenchyma. Therefore, regulation of deposition and assembly of secondary cell wall materials, i.e. phenolic components, appear to be disturbed by mutation of the cwa1 gene. Genetic analysis showed that cwa1 is allelic to brittle culm1 (bc1), which encodes the glycosylphosphatidylinositol-anchored COBRA-like protein specifically in plants. BC1 is known as a regulator that controls the culm mechanical strength and cellulose content in the secondary cell walls of sclerenchyma, but the precise function of BC1 has not been resolved. Our results suggest that CWA1/BC1 has an essential role in assembling cell wall constituents at their appropriate sites, thereby enabling synthesis of solid and flexible internodes in rice.

Cell Wall Remodeling by a Synthetic Analog Reveals Metabolic Adaptation in Vancomycin Resistant Enterococci.

PubMed

Pidgeon, Sean E; Pires, Marcos M

2017-07-21

Drug-resistant bacterial infections threaten to overburden our healthcare system and disrupt modern medicine. A large class of potent antibiotics, including vancomycin, operate by interfering with bacterial cell wall biosynthesis. Vancomycin-resistant enterococci (VRE) evade the blockage of cell wall biosynthesis by altering cell wall precursors, rendering them drug insensitive. Herein, we reveal the phenotypic plasticity and cell wall remodeling of VRE in response to vancomycin in live bacterial cells via a metabolic probe. A synthetic cell wall analog was designed and constructed to monitor cell wall structural alterations. Our results demonstrate that the biosynthetic pathway for vancomycin-resistant precursors can be hijacked by synthetic analogs to track the kinetics of phenotype induction. In addition, we leveraged this probe to interrogate the response of VRE cells to vancomycin analogs and a series of cell wall-targeted antibiotics. Finally, we describe a proof-of-principle strategy to visually inspect drug resistance induction. Based on our findings, we anticipate that our metabolic probe will play an important role in further elucidating the interplay among the enzymes involved in the VRE biosynthetic rewiring.

Do plant cell walls have a code?

PubMed

Tavares, Eveline Q P; Buckeridge, Marcos S

2015-12-01

A code is a set of rules that establish correspondence between two worlds, signs (consisting of encrypted information) and meaning (of the decrypted message). A third element, the adaptor, connects both worlds, assigning meaning to a code. We propose that a Glycomic Code exists in plant cell walls where signs are represented by monosaccharides and phenylpropanoids and meaning is cell wall architecture with its highly complex association of polymers. Cell wall biosynthetic mechanisms, structure, architecture and properties are addressed according to Code Biology perspective, focusing on how they oppose to cell wall deconstruction. Cell wall hydrolysis is mainly focused as a mechanism of decryption of the Glycomic Code. Evidence for encoded information in cell wall polymers fine structure is highlighted and the implications of the existence of the Glycomic Code are discussed. Aspects related to fine structure are responsible for polysaccharide packing and polymer-polymer interactions, affecting the final cell wall architecture. The question whether polymers assembly within a wall display similar properties as other biological macromolecules (i.e. proteins, DNA, histones) is addressed, i.e. do they display a code? Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria

PubMed Central

Mistou, Michel-Yves; Sutcliffe, Iain C.; van Sorge, Nina M.

2016-01-01

The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall teichoic acids (WTA). The presence and chemical structure of many rhamnose-containing cell wall polysaccharides (RhaCWP) has sometimes been known for decades. In contrast to WTA, insight into the biosynthesis and functional role of RhaCWP has been lacking. Recent studies in human streptococcal and enterococcal pathogens have highlighted critical roles for these complex polysaccharides in bacterial cell wall architecture and pathogenesis. In this review, we provide an overview of the RhaCWP with regards to their biosynthesis, genetics and biological function in species most relevant to human health. We also briefly discuss how increased knowledge in this field can provide interesting leads for new therapeutic compounds and improve biotechnological applications. PMID:26975195

Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria.

PubMed

Mistou, Michel-Yves; Sutcliffe, Iain C; van Sorge, Nina M

2016-07-01

The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall teichoic acids (WTA). The presence and chemical structure of many rhamnose-containing cell wall polysaccharides (RhaCWP) has sometimes been known for decades. In contrast to WTA, insight into the biosynthesis and functional role of RhaCWP has been lacking. Recent studies in human streptococcal and enterococcal pathogens have highlighted critical roles for these complex polysaccharides in bacterial cell wall architecture and pathogenesis. In this review, we provide an overview of the RhaCWP with regards to their biosynthesis, genetics and biological function in species most relevant to human health. We also briefly discuss how increased knowledge in this field can provide interesting leads for new therapeutic compounds and improve biotechnological applications. © FEMS 2016.

Synchrotron Time-Lapse Imaging of Lignocellulosic Biomass Hydrolysis: Tracking Enzyme Localization by Protein Autofluorescence and Biochemical Modification of Cell Walls by Microfluidic Infrared Microspectroscopy

PubMed Central

Devaux, Marie-Françoise; Jamme, Frédéric; André, William; Bouchet, Brigitte; Alvarado, Camille; Durand, Sylvie; Robert, Paul; Saulnier, Luc; Bonnin, Estelle; Guillon, Fabienne

2018-01-01

Tracking enzyme localization and following the local biochemical modification of the substrate should help explain the recalcitrance of lignocellulosic plant cell walls to enzymatic degradation. Time-lapse studies using conventional imaging require enzyme labeling and following the biochemical modifications of biopolymers found in plant cell walls, which cannot be easily achieved. In the present work, synchrotron facilities have been used to image the enzymatic degradation of lignocellulosic biomass without labeling the enzyme or the cell walls. Multichannel autofluorescence imaging of the protein and phenolic compounds after excitation at 275 nm highlighted the presence or absence of enzymes on cell walls and made it possible to track them during the reaction. Image analysis was used to quantify the fluorescence intensity variations. Consistent variations in the enzyme concentration were found locally for cell cavities and their surrounding cell walls. Microfluidic FT-IR microspectroscopy allowed for time-lapse tracking of local changes in the polysaccharides in cell walls during degradation. Hemicellulose degradation was found to occur prior to cellulose degradation using a Celluclast® preparation. Combining the fluorescence and FT-IR information yielded the conclusion that enzymes did not bind to lignified cell walls, which were consequently not degraded. Fluorescence multiscale imaging and FT-IR microspectroscopy showed an unexpected variability both in the initial biochemical composition and the degradation pattern, highlighting micro-domains in the cell wall of a given cell. Fluorescence intensity quantification showed that the enzymes were not evenly distributed, and their amount increased progressively on degradable cell walls. During degradation, adjacent cells were separated and the cell wall fragmented until complete degradation. PMID:29515611

Cell wall staining with Trypan blue enables quantitative analysis of morphological changes in yeast cells.

PubMed

Liesche, Johannes; Marek, Magdalena; Günther-Pomorski, Thomas

2015-01-01

Yeast cells are protected by a cell wall that plays an important role in the exchange of substances with the environment. The cell wall structure is dynamic and can adapt to different physiological states or environmental conditions. For the investigation of morphological changes, selective staining with fluorescent dyes is a valuable tool. Furthermore, cell wall staining is used to facilitate sub-cellular localization experiments with fluorescently-labeled proteins and the detection of yeast cells in non-fungal host tissues. Here, we report staining of Saccharomyces cerevisiae cell wall with Trypan Blue, which emits strong red fluorescence upon binding to chitin and yeast glucan; thereby, it facilitates cell wall analysis by confocal and super-resolution microscopy. The staining pattern of Trypan Blue was similar to that of the widely used UV-excitable, blue fluorescent cell wall stain Calcofluor White. Trypan Blue staining facilitated quantification of cell size and cell wall volume when utilizing the optical sectioning capacity of a confocal microscope. This enabled the quantification of morphological changes during growth under anaerobic conditions and in the presence of chemicals, demonstrating the potential of this approach for morphological investigations or screening assays.

A unified wall function for compressible turbulence modelling

NASA Astrophysics Data System (ADS)

Ong, K. C.; Chan, A.

2018-05-01

Turbulence modelling near the wall often requires a high mesh density clustered around the wall and the first cells adjacent to the wall to be placed in the viscous sublayer. As a result, the numerical stability is constrained by the smallest cell size and hence requires high computational overhead. In the present study, a unified wall function is developed which is valid for viscous sublayer, buffer sublayer and inertial sublayer, as well as including effects of compressibility, heat transfer and pressure gradient. The resulting wall function applies to compressible turbulence modelling for both isothermal and adiabatic wall boundary conditions with the non-zero pressure gradient. Two simple wall function algorithms are implemented for practical computation of isothermal and adiabatic wall boundary conditions. The numerical results show that the wall function evaluates the wall shear stress and turbulent quantities of wall adjacent cells at wide range of non-dimensional wall distance and alleviate the number and size of cells required.

Growth and cell wall changes in stem organs under microgravity and hypergravity conditions

NASA Astrophysics Data System (ADS)

Hoson, Takayuki; Soga, Kouichi; Wakabayashi, Kazuyuki; Kamisaka, Seiichiro

Gravity strongly influences plant growth and development, which is fundamentally brought about by modifications to the properties of the cell wall. We have examined the changes in growth and cell wall properties in seedling organs under hypergravity conditions produced by centrifugation and under microgravity conditions in space. Hypergravity stimuli have been shown to decrease the growth rate of various seedling organs. When hypergravity suppressed elongation growth, a decrease in cell wall extensibility (an increase in cell wall rigidity) was induced. Hypergravity has also been shown to increase cell wall thickness in various mate-rials. In addition, a polymerization of certain matrix polysaccharides was brought about by hypergravity: in dicotyledons hypergravity increased the molecular size of xyloglucans, whereas hypergravity increased that of 1,3,1,4-β-glucans in monocotyledonous Gramineae. These mod-ifications to cell wall metabolism may be responsible for a decrease in cell wall extensibility, leading to growth suppression under hypergravity conditions. How then does microgravity in-fluence growth and cell wall properties? Here, there was a possibility that microgravity might induce changes similar to those by hypergravity, because plants have evolved and adapted to 1 g condition for more than 400 million years. However, the changes observed under microgravity conditions in space were just opposite to those induced by hypergravity: stimulation of elonga-tion growth, an increase in cell wall extensibility, and a decrease in cell wall thickness as well as depolymerization of cell wall polysaccharides were brought about in space. Furthermore, growth and cell wall properties varied in proportion to the logarithm of the magnitude of grav-ity in the range from microgravity to hypergravity, as shown in the dose-response relation in light and hormonal responses. Thus, microgravity may be a `stress-less' environment for plant seedlings to grow and develop. Preliminary results obtained by recent Space Seed experiment in the Kibo Module on the International Space Station (PI: S. Kamisaka) suggest that this hypothesis is also applicable to mature Arabidopsis plants.

Profiling the Hydrolysis of Isolated Grape Berry Skin Cell Walls by Purified Enzymes.

PubMed

Zietsman, Anscha J J; Moore, John P; Fangel, Jonatan U; Willats, William G T; Vivier, Melané A

2015-09-23

The unraveling of crushed grapes by maceration enzymes during winemaking is difficult to study because of the complex and rather undefined nature of both the substrate and the enzyme preparations. In this study we simplified both the substrate, by using isolated grape skin cell walls, and the enzyme preparations, by using purified enzymes in buffered conditions, to carefully follow the impact of the individual and combined enzymes on the grape skin cell walls. By using cell wall profiling techniques we could monitor the compositional changes in the grape cell wall polymers due to enzyme activity. Extensive enzymatic hydrolysis, achieved with a preparation of pectinases or pectinases combined with cellulase or hemicellulase enzymes, completely removed or drastically reduced levels of pectin polymers, whereas less extensive hydrolysis only opened up the cell wall structure and allowed extraction of polymers from within the cell wall layers. Synergistic enzyme activity was detectable as well as indications of specific cell wall polymer associations.

Outside-in control -Does plant cell wall integrity regulate cell cycle progression?

PubMed

Gigli-Bisceglia, Nora; Hamann, Thorsten

2018-04-13

During recent years it has become accepted that plant cell walls are not inert objects surrounding all plant cells but are instead highly dynamic, plastic structures. They are involved in a large number of cell biological processes and contribute actively to plant growth, development and interaction with environment. Therefore, it is not surprising that cellular processes can control plant cell wall integrity while, simultaneously, cell wall integrity can influence cellular processes. In yeast and animal cells such a bi-directional relationship also exists between the yeast/animal extra-cellular matrices and the cell cycle. In yeast, the cell wall integrity maintenance mechanism and a dedicated plasmamembrane integrity checkpoint are mediating this relationship. Recent research has yielded insights into the mechanism controlling plant cell wall metabolism during cytokinesis. However, knowledge regarding putative regulatory pathways controlling adaptive modifications in plant cell cycle activity in response to changes in the state of the plant cell wall are not yet identified. In this review, we summarize similarities and differences in regulatory mechanisms coordinating extra cellular matrices and cell cycle activity in animal and yeast cells, discuss the available evidence supporting the existence of such a mechanism in plants and suggest that the plant cell wall integrity maintenance mechanism might also control cell cycle activity in plant cells. This article is protected by copyright. All rights reserved.

Phenotype-Based Screening of Small Molecules to Modify Plant Cell Walls Using BY-2 Cells.

PubMed

Okubo-Kurihara, Emiko; Matsui, Minami

2018-01-01

The plant cell wall is an important and abundant biomass with great potential for use as a modern recyclable resource. For effective utilization of this cellulosic biomass, its ability to degrade efficiently is key point. With the aim of modifying the cell wall to allow easy decomposition, we used chemical biological technology to alter its structure. As a first step toward evaluating the chemicals in the cell wall we employed a phenotype-based approach using high-throughput screening. As the plant cell wall is essential in determining cell morphology, phenotype-based screening is particularly effective in identifying compounds that bring about alterations in the cell wall. For rapid and reproducible screening, tobacco BY-2 cell is an excellent system in which to observe cell morphology. In this chapter, we provide a detailed chemical biological methodology for studying cell morphology using tobacco BY-2 cells.

The Genus Cladophora Kützing (Ulvophyceae) as a Globally Distributed Ecological Engineer.

PubMed

Zulkifly, Shahrizim B; Graham, James M; Young, Erica B; Mayer, Robert J; Piotrowski, Michael J; Smith, Izak; Graham, Linda E

2013-02-01

The green algal genus Cladophora forms conspicuous nearshore populations in marine and freshwaters worldwide, commonly dominating peri-phyton communities. As the result of human activities, including the nutrient pollution of nearshore waters, Cladophora-dominated periphyton can form nuisance blooms. On the other hand, Cladophora has ecological functions that are beneficial, but less well appreciated. For example, Cladophora has previously been characterized as an ecological engineer because its complex structure fosters functional and taxonomic diversity of benthic microfauna. Here, we review classic and recent literature concerning taxonomy, cell biology, morphology, reproductive biology, and ecology of the genus Cladophora, to examine how this alga functions to modify habitats and influence littoral biogeochemistry. We review the evidence that Cladophora supports large, diverse populations of microalgal and bacterial epiphytes that influence the cycling of carbon and other key elements, and that the high production of cellulose and hydrocarbons by Cladophora-dominated periphyton has the potential for diverse technological applications, including wastewater remediation coupled to renewable biofuel production. We postulate that well-known aspects of Cladophora morphology, hydrodynamically stable and perennial holdfasts, distinctively branched architecture, unusually large cell and sporangial size and robust cell wall construction, are major factors contributing to the multiple roles of this organism as an ecological engineer. © 2013 Phycological Society of America.

The plant cell wall in the feeding sites of cyst nematodes.

PubMed

Bohlmann, Holger; Sobczak, Miroslaw

2014-01-01

Plant parasitic cyst nematodes (genera Heterodera and Globodera) are serious pests for many crops. They enter the host roots as migratory second stage juveniles (J2) and migrate intracellularly toward the vascular cylinder using their stylet and a set of cell wall degrading enzymes produced in the pharyngeal glands. They select an initial syncytial cell (ISC) within the vascular cylinder or inner cortex layers to induce the formation of a multicellular feeding site called a syncytium, which is the only source of nutrients for the parasite during its entire life. A syncytium can consist of more than hundred cells whose protoplasts are fused together through local cell wall dissolutions. While the nematode produces a cocktail of cell wall degrading and modifying enzymes during migration through the root, the cell wall degradations occurring during syncytium development are due to the plants own cell wall modifying and degrading proteins. The outer syncytial cell wall thickens to withstand the increasing osmotic pressure inside the syncytium. Furthermore, pronounced cell wall ingrowths can be formed on the outer syncytial wall at the interface with xylem vessels. They increase the surface of the symplast-apoplast interface, thus enhancing nutrient uptake into the syncytium. Processes of cell wall degradation, synthesis and modification in the syncytium are facilitated by a variety of plant proteins and enzymes including expansins, glucanases, pectate lyases and cellulose synthases, which are produced inside the syncytium or in cells surrounding the syncytium.

Cell wall of pathogenic yeasts and implications for antimycotic therapy.

PubMed

Cassone, A

1986-01-01

Yeast cell wall is a complex, multilayered structure where amorphous, granular and fibrillar components interact with each other to confer both the specific cell shape and osmotic protection against lysis. Thus it is widely recognized that as is the case with bacteria, yeast cell wall is a major potential target for selective chemotherapeutic drugs. Despite intensive research, very few such drugs have been discovered and none has found substantial application in human diseases to date. Among the different cell wall components, beta-glucan and chitin are the fibrillar materials playing a fundamental role in the overall rigidity and resistance of the wall. Inhibition of the metabolism of these polymers, therefore, should promptly lead to lysis. This indeed occurs and aculeacin, echinocandin and polyoxins are examples of agents producing such an action. Particular attention should be focused on chitin synthesis. Although quantitatively a minor cell wall component, chitin is important in the mechanism of dimorphic transition, especially in Candida albicans, a major human opportunistic pathogen. This transition is associated with increased invasiveness and general virulence of the fungus. Yeast cell wall may also limit the effect of antifungals which owe their action to disturbance of the cytoplasmic membrane or of cell metabolism. Indeed, the cell wall may hinder access to the cell interior both under growing conditions and, particularly, during cell ageing in the stationary phase, when important structural changes occur in the cell wall due to unbalanced wall growth (phenotypic drug resistance).

An Analysis of Insulated Concrete Forms for use in Sustainable Military Construction

DTIC Science & Technology

2014-03-27

CONSTRUCTION THESIS Presented to the Faculty Department of Systems and Engineering Management Graduate School of Engineering and Management...which fit together and are filled with reinforced concrete to construct the exterior wall systems of a building. By design, this material provides a...Forms with Rebar .............................................................. 12 Figure 3. Cut outs of ICF wall systems

Development of Cellulosic Biofuels (LBNL Summer Lecture Series)

ScienceCinema

Somerville, Chris [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Biosciences Division; Stanford Univ., CA (United States). Dept. of Biological Sciences

2018-05-18

Summer Lecture Series 2007: Chris Somerville, Director of the Energy Biosciences Institute and an award-winning plant biochemist with Berkeley Lab's Physical Biosciences Division, is a leading authority on the structure and function of plant cell walls. He discusses an overview of some of the technical challenges associated with the production of cellulosic biofuels, which will require an improved understanding of a diverse range of topics in fields such as agronomy, chemical engineering, microbiology, structural biology, genomics, environmental sciences, and socioeconomics.

Engineering Assessment of Big House at Summit Station, Greenland

DTIC Science & Technology

2011-04-01

Open cell expanded polystyrene insulation c EPS is not thought to be under the T-111 siding (communication 04 Nov 2010) d Quad-pane window ERDC...eastern side of the building where, in 2008, the roof access hatch was relocated. In 1999, the exterior was re-sheathed with 38- mm expanded ... polystyrene (EPS) insulation and 12.7-mm T-111 siding, which was attached with a number of fasteners. The wall thickness is 178 mm (Fig. 3). In 2006, a

A&M. Hot liquid waste treatment building (TAN616). Contextual view, facing ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

A&M. Hot liquid waste treatment building (TAN-616). Contextual view, facing south. Wall of hot shop (TAN-607) with high bay at left of view. Lower-roofed building at left edge of view is TAN- 633, hot cell annex. Complex at center of view is TAN-616. Tall metal building with gable roof is TAN-615. Photographer: Ron Paarmann. Date: September 22, 1997. INEEL negative no. HD-20-2-2 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

Preconstruction Engineering and Design Phase Reevaluation Report, Olcott Harbor Project, Olcott, New York. Volume 2, Supporting Documentation. Revision.

DTIC Science & Technology

1992-09-01

85 (saturated) : 0 : 26 3. Cell Fill (crushed stone) : 110 (moist) : 0 : 32 68 ( submerged ) 4. Breakwater Armor Stone : 110 (moist) : 0 40 68... submerged ) 5. Queenston Shale bedrock : unconfined compressive strength : au = 6,000 psi 7 5. PROJECT FEATURES FOR FINAL DESIGN a. Breakwaters. The project...existing conrete pad (550 feet from the retaining wall) the structure is submerged with isolated portions of the structure observable at the lake

Plant cell wall extensibility: connecting plant cell growth with cell wall structure, mechanics, and the action of wall-modifying enzymes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cosgrove, Daniel J.

The advent of user-friendly instruments for measuring force/deflection curves of plant surfaces at high spatial resolution has resulted in a recent outpouring of reports of the ‘Young's modulus’ of plant cell walls. The stimulus for these mechanical measurements comes from biomechanical models of morphogenesis of meristems and other tissues, as well as single cells, in which cell wall stress feeds back to regulate microtubule organization, auxin transport, cellulose deposition, and future growth directionality. In this article I review the differences between elastic modulus and wall extensibility in the context of cell growth. Some of the inherent complexities, assumptions, and potentialmore » pitfalls in the interpretation of indentation force/deflection curves are discussed. Reported values of elastic moduli from surface indentation measurements appear to be 10- to >1000-fold smaller than realistic tensile elastic moduli in the plane of plant cell walls. Potential reasons for this disparity are discussed, but further work is needed to make sense of the huge range in reported values. The significance of wall stress relaxation for growth is reviewed and connected to recent advances and remaining enigmas in our concepts of how cellulose, hemicellulose, and pectins are assembled to make an extensible cell wall. A comparison of the loosening action of α-expansin and Cel12A endoglucanase is used to illustrate two different ways in which cell walls may be made more extensible and the divergent effects on wall mechanics.« less

My body is a cage: mechanisms and modulation of plant cell growth.

PubMed

Braidwood, Luke; Breuer, Christian; Sugimoto, Keiko

2014-01-01

388 I. 388 II. 389 III. 389 IV. 390 V. 391 VI. 393 VII. 394 VIII. 398 399 References 399 SUMMARY: The wall surrounding plant cells provides protection from abiotic and biotic stresses, and support through the action of turgor pressure. However, the presence of this strong elastic wall also prevents cell movement and resists cell growth. This growth can be likened to extending a house from the inside, using extremely high pressures to push out the walls. Plants must increase cell volume in order to explore their environment, acquire nutrients and reproduce. Cell wall material must stretch and flow in a controlled manner and, concomitantly, new cell wall material must be deposited at the correct rate and site to prevent wall and cell rupture. In this review, we examine biomechanics, cell wall structure and growth regulatory networks to provide a 'big picture' of plant cell growth. © 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.

The connection of cytoskeletal network with plasma membrane and the cell wall

PubMed Central

Liu, Zengyu; Persson, Staffan; Zhang, Yi

2015-01-01

The cell wall provides external support of the plant cells, while the cytoskeletons including the microtubules and the actin filaments constitute an internal framework. The cytoskeletons contribute to the cell wall biosynthesis by spatially and temporarily regulating the transportation and deposition of cell wall components. This tight control is achieved by the dynamic behavior of the cytoskeletons, but also through the tethering of these structures to the plasma membrane. This tethering may also extend beyond the plasma membrane and impact on the cell wall, possibly in the form of a feedback loop. In this review, we discuss the linking components between the cytoskeletons and the plasma membrane, and/or the cell wall. We also discuss the prospective roles of these components in cell wall biosynthesis and modifications, and aim to provide a platform for further studies in this field. PMID:25693826

Tuned intake air system for a rotary engine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Corbett, W.D.

This patent describes a rotary internal combustion engine for an outboard board motor. It comprises a plenum chamber attached to the rear of the engine; and the plenum chamber including an inner wall attached to the exhaust manifold; an inlet conduit connecting the cooling air exit passage and the inlet air opening; an outlet conduit connecting the outlet air opening and the combustion air inlet; and the outlet conduit terminating in a combustion air outlet in the inner wall of the plenum chamber.

Cellulose synthase complexes display distinct dynamic behaviors during xylem transdifferentiation.

PubMed

Watanabe, Yoichiro; Schneider, Rene; Barkwill, Sarah; Gonzales-Vigil, Eliana; Hill, Joseph L; Samuels, A Lacey; Persson, Staffan; Mansfield, Shawn D

2018-06-05

In plants, plasma membrane-embedded CELLULOSE SYNTHASE (CESA) enzyme complexes deposit cellulose polymers into the developing cell wall. Cellulose synthesis requires two different sets of CESA complexes that are active during cell expansion and secondary cell wall thickening, respectively. Hence, developing xylem cells, which first undergo cell expansion and subsequently deposit thick secondary walls, need to completely reorganize their CESA complexes from primary wall- to secondary wall-specific CESAs. Using live-cell imaging, we analyzed the principles underlying this remodeling. At the onset of secondary wall synthesis, the primary wall CESAs ceased to be delivered to the plasma membrane and were gradually removed from both the plasma membrane and the Golgi. For a brief transition period, both primary wall- and secondary wall-specific CESAs coexisted in banded domains of the plasma membrane where secondary wall synthesis is concentrated. During this transition, primary and secondary wall CESAs displayed discrete dynamic behaviors and sensitivities to the inhibitor isoxaben. As secondary wall-specific CESAs were delivered and inserted into the plasma membrane, the primary wall CESAs became concentrated in prevacuolar compartments and lytic vacuoles. This adjustment in localization between the two CESAs was accompanied by concurrent decreased primary wall CESA and increased secondary wall CESA protein abundance. Our data reveal distinct and dynamic subcellular trafficking patterns that underpin the remodeling of the cellulose biosynthetic machinery, resulting in the removal and degradation of the primary wall CESA complex with concurrent production and recycling of the secondary wall CESAs. Copyright © 2018 the Author(s). Published by PNAS.

Numerical modelling of closed-cell aluminium foam under dynamic loading

NASA Astrophysics Data System (ADS)

Hazell, Paul; Kader, M. A.; Islam, M. A.; Escobedo, J. P.; Saadatfar, M.

2015-06-01

Closed-cell aluminium foams are extensively used in aerospace and automobile industries. The understanding of their behaviour under impact loading conditions is extremely important since impact problems are directly related to design of these engineering structures. This research investigates the response of a closed-cell aluminium foam (CYMAT) subjected to dynamic loading using the finite element software ABAQUS/explicit. The aim of this research is to numerically investigate the material and structural properties of closed-cell aluminium foam under impact loading conditions with interest in shock propagation and its effects on cell wall deformation. A μ-CT based 3D foam geometry is developed to simulate the local cell collapse behaviours. A number of numerical techniques are applied for modelling the crush behaviour of aluminium foam to obtain the more accurate results. The simulation results are compared with experimental data. Comparison of the results shows a good correlation between the experimental results and numerical predictions.

Identification of potential cell wall component that allows Taka-amylase A adsorption in submerged cultures of Aspergillus oryzae.

PubMed

Sato, Hiroki; Toyoshima, Yoshiyuki; Shintani, Takahiro; Gomi, Katsuya

2011-12-01

We observed that α-amylase (Taka-amylase A; TAA) activity in the culture broth disappeared in the later stage of submerged cultivation of Aspergillus oryzae. This disappearance was caused by adsorption of TAA onto the cell wall of A. oryzae and not due to protein degradation by extracellular proteolytic enzymes. To determine the cell wall component(s) that allows TAA adsorption efficiently, the cell wall was fractionated by stepwise alkali treatment and enzymatic digestion. Consequently, alkali-insoluble cell wall fractions exhibited high levels of TAA adsorption. In addition, this adsorption capacity was significantly enhanced by treatment of the alkali-insoluble fraction with β-glucanase, which resulted in the concomitant increase in the amount of chitin in the resulting fraction. In contrast, the adsorption capacity was diminished by treating the cell wall fraction with chitinase. These results suggest that the major component that allows TAA adsorption is chitin. However, both the mycelium and the cell wall demonstrated the inability to allow TAA adsorption in the early stage of cultivation, despite chitin content in the cell wall being identical in both early and late stages of cultivation. These results suggest the existence of unidentified factor(s) that could prevent the adsorption of TAA onto the cell wall. Such factor(s) is most likely removed or diminished from the cell wall following longer cultivation periods.

A model of cell wall expansion based on thermodynamics of polymer networks

NASA Technical Reports Server (NTRS)

Veytsman, B. A.; Cosgrove, D. J.

1998-01-01

A theory of cell wall extension is proposed. It is shown that macroscopic properties of cell walls can be explained through the microscopic properties of interpenetrating networks of cellulose and hemicellulose. The qualitative conclusions of the theory agree with the existing experimental data. The dependence of the cell wall yield threshold on the secretion of the wall components is discussed.

Bacterial cell wall composition and the influence of antibiotics by cell-wall and whole-cell NMR

PubMed Central

Romaniuk, Joseph A. H.; Cegelski, Lynette

2015-01-01

The ability to characterize bacterial cell-wall composition and structure is crucial to understanding the function of the bacterial cell wall, determining drug modes of action and developing new-generation therapeutics. Solid-state NMR has emerged as a powerful tool to quantify chemical composition and to map cell-wall architecture in bacteria and plants, even in the context of unperturbed intact whole cells. In this review, we discuss solid-state NMR approaches to define peptidoglycan composition and to characterize the modes of action of old and new antibiotics, focusing on examples in Staphylococcus aureus. We provide perspectives regarding the selected NMR strategies as we describe the exciting and still-developing cell-wall and whole-cell NMR toolkit. We also discuss specific discoveries regarding the modes of action of vancomycin analogues, including oritavancin, and briefly address the reconsideration of the killing action of β-lactam antibiotics. In such chemical genetics approaches, there is still much to be learned from perturbations enacted by cell-wall assembly inhibitors, and solid-state NMR approaches are poised to address questions of cell-wall composition and assembly in S. aureus and other organisms. PMID:26370936

A Comparative Study of Sample Preparation for Staining and Immunodetection of Plant Cell Walls by Light Microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Verhertbruggen, Yves; Walker, Jesse L.; Guillon, Fabienne

Staining and immunodetection by light microscopy are methods widely used to investigate plant cell walls. The two techniques have been crucial to study the cell wall architecture in planta, its deconstruction by chemicals or cell wall-degrading enzymes. They have been instrumental in detecting the presence of cell types, in deciphering plant cell wall evolution and in characterizing plant mutants and transformants. The success of immunolabeling relies on how plant materials are embedded and sectioned. Agarose coating, wax and resin embedding are, respectively, associated with vibratome, microtome and ultramicrotome sectioning. Here, we have systematically carried out a comparative analysis of thesemore » three methods of sample preparation when they are applied for cell wall staining and cell wall immunomicroscopy. In order to help the plant community in understanding and selecting adequate methods of embedding and sectioning for cell wall immunodetection, we review in this article the advantages and limitations of these three methods. Moreover, we offer detailed protocols of embedding for studying plant materials through microscopy.« less

A Comparative Study of Sample Preparation for Staining and Immunodetection of Plant Cell Walls by Light Microscopy

DOE PAGES

Verhertbruggen, Yves; Walker, Jesse L.; Guillon, Fabienne; ...

2017-08-29

Staining and immunodetection by light microscopy are methods widely used to investigate plant cell walls. The two techniques have been crucial to study the cell wall architecture in planta, its deconstruction by chemicals or cell wall-degrading enzymes. They have been instrumental in detecting the presence of cell types, in deciphering plant cell wall evolution and in characterizing plant mutants and transformants. The success of immunolabeling relies on how plant materials are embedded and sectioned. Agarose coating, wax and resin embedding are, respectively, associated with vibratome, microtome and ultramicrotome sectioning. Here, we have systematically carried out a comparative analysis of thesemore » three methods of sample preparation when they are applied for cell wall staining and cell wall immunomicroscopy. In order to help the plant community in understanding and selecting adequate methods of embedding and sectioning for cell wall immunodetection, we review in this article the advantages and limitations of these three methods. Moreover, we offer detailed protocols of embedding for studying plant materials through microscopy.« less

A Comparative Study of Sample Preparation for Staining and Immunodetection of Plant Cell Walls by Light Microscopy

PubMed Central

Verhertbruggen, Yves; Walker, Jesse L.; Guillon, Fabienne; Scheller, Henrik V.

2017-01-01

Staining and immunodetection by light microscopy are methods widely used to investigate plant cell walls. The two techniques have been crucial to study the cell wall architecture in planta, its deconstruction by chemicals or cell wall-degrading enzymes. They have been instrumental in detecting the presence of cell types, in deciphering plant cell wall evolution and in characterizing plant mutants and transformants. The success of immunolabeling relies on how plant materials are embedded and sectioned. Agarose coating, wax and resin embedding are, respectively, associated with vibratome, microtome and ultramicrotome sectioning. Here, we have systematically carried out a comparative analysis of these three methods of sample preparation when they are applied for cell wall staining and cell wall immunomicroscopy. In order to help the plant community in understanding and selecting adequate methods of embedding and sectioning for cell wall immunodetection, we review in this article the advantages and limitations of these three methods. Moreover, we offer detailed protocols of embedding for studying plant materials through microscopy. PMID:28900439

Combustor nozzles in gas turbine engines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnson, Thomas Edward; Keener, Christopher Paul; Stewart, Jason Thurman

2017-09-12

A micro-mixer nozzle for use in a combustor of a combustion turbine engine, the micro-mixer nozzle including: a fuel plenum defined by a shroud wall connecting a periphery of a forward tube sheet to a periphery of an aft tubesheet; a plurality of mixing tubes extending across the fuel plenum for mixing a supply of compressed air and fuel, each of the mixing tubes forming a passageway between an inlet formed through the forward tubesheet and an outlet formed through the aft tubesheet; and a wall mixing tube formed in the shroud wall.

Microrheological studies reveal semiflexible networks in gels of a ubiquitous cell wall polysaccharide

NASA Astrophysics Data System (ADS)

Vincent, R. R.; Pinder, D. N.; Hemar, Y.; Williams, M. A. K.

2007-09-01

Microrheological measurements have been carried out on ionotropic gels made from an important cell wall polysaccharide, using diffusing wave spectroscopy and multiple particle tracking. These gels were formed by the interaction of calcium ions with negatively charged groups on the polymer backbone, which is a copolymer of charged and uncharged sugars, galacturonic acid, and its methylesterified analog, respectively. The results suggest that semiflexible networks are formed in these systems, with a low frequency, frequency independent storage modulus (G'>G″) , and a high frequency scaling of both G' and G″ with ω3/4 . The differences observed between gels obtained using polysaccharide samples with different amounts and patterns of the charged ion-binding groups could comfortably be accommodated within this theoretical framework, assuming that the elementary semiflexible elements of the network are filaments consisting of two polymer chains bridged with calcium. In particular, a sample that was engineered to possess a blockwise intramolecular distribution of calcium chelating moieties clearly exhibited the high frequency scaling of both moduli with ω3/4 across some three orders of magnitude, and the concentration dependences of the elastic modulus, at both high and low frequency, were found to follow power laws with predicted exponents. Furthermore, quantitative agreement of the moduli with theory was found for realistic estimates of the molecular parameters, suggesting that the physics of semiflexible networks is not only exploited by protein components of the cytoskeleton but also by polysaccharides in plant cell walls.

Bioinspired assemblies of plant cell wall polymers unravel the affinity properties of carbohydrate-binding modules.

PubMed

Paës, Gabriel; von Schantz, Laura; Ohlin, Mats

2015-09-07

Lignocellulose-acting enzymes play a central role in the biorefinery of plant biomass to make fuels, chemicals and materials. These enzymes are often appended to carbohydrate binding modules (CBMs) that promote substrate targeting. When used in plant materials, which are complex assemblies of polymers, the binding properties of CBMs can be difficult to understand and predict, thus limiting the efficiency of enzymes. In order to gain more information on the binding properties of CBMs, some bioinspired model assemblies that contain some of the polymers and covalent interactions found in the plant cell walls have been designed. The mobility of three engineered CBMs has been investigated by FRAP in these assemblies, while varying the parameters related to the polymer concentration, the physical state of assemblies and the oligomerization state of CBMs. The features controlling the mobility of the CBMs in the assemblies have been quantified and hierarchized. We demonstrate that the parameters can have additional or opposite effects on mobility, depending on the CBM tested. We also find evidence of a relationship between the mobility of CBMs and their binding strength. Overall, bioinspired assemblies are able to reveal the unique features of affinity of CBMs. In particular, the results show that oligomerization of CBMs and the presence of ferulic acid motifs in the assemblies play an important role in the binding affinity of CBMs. Thus we propose that these features should be finely tuned when CBMs are used in plant cell walls to optimise bioprocesses.

Comparative chemical characterization of pigmented and less pigmented cell walls of Alternaria tenuissima.

PubMed

Kishore, Kankipati Hara; Kanjilal, Sanjit; Misra, Sunil; Reddy, Chinnathimma Rajagopal; Murty, Upadyayula Suryanarayana

2005-12-01

Alternaria tenuissima, the parasitic fungus, was obtained from the pruned upper-cut surfaces of mulberry stems. This fungus contains dark pigment because of the presence of melanin in the cell wall. To obtain less-pigmented cell walls, this fungus was grown under dark condition. When the pigmented and less-pigmented cell walls were chemically analyzed, no differences were observed in amino-acid composition, hexoses, or pentoses. However, in pigmented cell walls, higher contents of melanin (2.6%) were found than in less-pigmented cell walls (0.3%). Interestingly, a significant difference was observed in the relative fatty-acid compositions between these two types of cell walls. Among the major fatty acids, there were increased concentrations of tetradecanoic acid (C14:0), hexadecanoic acid (C16:0), 9-hexadecenoic acid (C16: 1,Delta 9), and 9-octadecanoic acid (C18:1,Delta 9) and a concomitant decrease in 9,12-octadecadienoic acid (C18:2,Delta 9,12) in less-pigmented compared with pigmented cell walls. This difference in fatty-acid composition may be related to the higher percentage of melanin in the pigmented than the less-pigmented cell walls. Lesser amounts of 9,12-octadecadienoic acid in less-pigmented cell walls may have been caused by the growth of the fungus under environmental stress conditions. An interesting observation was the presence in pigmented cell walls only of methyl-substituted fatty acids with carbon numbers C14 to C17, but their occurrence could not be ascertained in the present study.

Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption

PubMed Central

Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

2015-01-01

The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry. PMID:26295574

Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption.

PubMed

Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

2015-01-01

The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry.

Agronomic performance of Populus deltoides trees engineered for biofuel production

DOE PAGES

Macaya-Sanz, David; Chen, Jin?Gui; Kalluri, Udaya C.; ...

2017-11-30

Background: One of the major barriers to the development of lignocellulosic feedstocks is the recalcitrance of plant cell walls to deconstruction and saccharification. Recalcitrance can be reduced by targeting genes involved in cell wall biosynthesis, but this can have unintended consequences that compromise the agronomic performance of the trees under field conditions. Here we report the results of a field trial of fourteen distinct transgenic Populus deltoides lines that had previously demonstrated reduced recalcitrance without yield penalties under greenhouse conditions.Results: Survival and productivity of the trial were excellent in the first year, and there was little evidence for reduced performancemore » of the transgenic lines with modified target gene expression. Surprisingly, the most striking phenotypic effects in this trial were for two empty-vector control lines that had modified bud set and bud flush. This is most likely due to somaclonal variation or insertional mutagenesis. Traits related to yield, crown architecture, herbivory, pathogen response, and frost damage showed few significant differences between target gene transgenics and empty vector controls. However, there were a few interesting exceptions. Lines overexpressing the DUF231 gene, a putative O-acetyltransferase, showed early bud flush and marginally increased height growth. Lines overexpressing the DUF266 gene, a putative glycosyltransferase, had significantly decreased stem internode length and slightly higher volume index. Finally, lines overexpressing the PFD2 gene, a putative member of the prefoldin complex, had a slightly reduced volume index.Conclusions: This field trial demonstrates that these cell wall modifications, which decreased cell wall recalcitrance under laboratory conditions, did not seriously compromise first-year performance in the field, despite substantial challenges, including an outbreak of a stem boring insect (Gypsonoma haimbachiana), attack by a leaf rust pathogen (Melampsora spp.), and a late frost event. This bodes well for the potential utility of these lines as advanced biofuels feedstocks.« less

Agronomic performance of Populus deltoides trees engineered for biofuel production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Macaya-Sanz, David; Chen, Jin?Gui; Kalluri, Udaya C.

Background: One of the major barriers to the development of lignocellulosic feedstocks is the recalcitrance of plant cell walls to deconstruction and saccharification. Recalcitrance can be reduced by targeting genes involved in cell wall biosynthesis, but this can have unintended consequences that compromise the agronomic performance of the trees under field conditions. Here we report the results of a field trial of fourteen distinct transgenic Populus deltoides lines that had previously demonstrated reduced recalcitrance without yield penalties under greenhouse conditions.Results: Survival and productivity of the trial were excellent in the first year, and there was little evidence for reduced performancemore » of the transgenic lines with modified target gene expression. Surprisingly, the most striking phenotypic effects in this trial were for two empty-vector control lines that had modified bud set and bud flush. This is most likely due to somaclonal variation or insertional mutagenesis. Traits related to yield, crown architecture, herbivory, pathogen response, and frost damage showed few significant differences between target gene transgenics and empty vector controls. However, there were a few interesting exceptions. Lines overexpressing the DUF231 gene, a putative O-acetyltransferase, showed early bud flush and marginally increased height growth. Lines overexpressing the DUF266 gene, a putative glycosyltransferase, had significantly decreased stem internode length and slightly higher volume index. Finally, lines overexpressing the PFD2 gene, a putative member of the prefoldin complex, had a slightly reduced volume index.Conclusions: This field trial demonstrates that these cell wall modifications, which decreased cell wall recalcitrance under laboratory conditions, did not seriously compromise first-year performance in the field, despite substantial challenges, including an outbreak of a stem boring insect (Gypsonoma haimbachiana), attack by a leaf rust pathogen (Melampsora spp.), and a late frost event. This bodes well for the potential utility of these lines as advanced biofuels feedstocks.« less

Mating-Induced Shedding of Cell Walls, Removal of Walls from Vegetative Cells, and Osmotic Stress Induce Presumed Cell Wall Genes in Chlamydomonas1

PubMed Central

Hoffmann, Xenia-Katharina; Beck, Christoph F.

2005-01-01

The first step in sexual differentiation of the unicellular green alga Chlamydomonas reinhardtii is the formation of gametes. Three genes, GAS28, GAS30, and GAS31, encoding Hyp-rich glycoproteins that presumably are cell wall constituents, are expressed in the late phase of gametogenesis. These genes, in addition, are activated by zygote formation and cell wall removal and by the application of osmotic stress. The induction by zygote formation could be traced to cell wall shedding prior to gamete fusion since it was seen in mutants defective in cell fusion. However, it was absent in mutants defective in the initial steps of mating, i.e. in flagellar agglutination and in accumulation of adenosine 3′,5′-cyclic monophosphate in response to this agglutination. Induction of the three GAS genes was also observed when cultures were exposed to hypoosmotic or hyperosmotic stress. To address the question whether the induction seen upon cell wall removal from both gametes and vegetative cells was elicited by osmotic stress, cell wall removal was performed under isosmotic conditions. Also under such conditions an activation of the genes was observed, suggesting that the signaling pathway(s) is (are) activated by wall removal itself. PMID:16183845

Discovery of Novel Cell Wall-Active Compounds Using PywaC, a Sensitive Reporter of Cell Wall Stress, in the Model Gram-Positive Bacterium Bacillus subtilis

PubMed Central

Czarny, T. L.; Perri, A. L.; French, S.

2014-01-01

The emergence of antibiotic resistance in recent years has radically reduced the clinical efficacy of many antibacterial treatments and now poses a significant threat to public health. One of the earliest studied well-validated targets for antimicrobial discovery is the bacterial cell wall. The essential nature of this pathway, its conservation among bacterial pathogens, and its absence in human biology have made cell wall synthesis an attractive pathway for new antibiotic drug discovery. Herein, we describe a highly sensitive screening methodology for identifying chemical agents that perturb cell wall synthesis, using the model of the Gram-positive bacterium Bacillus subtilis. We report on a cell-based pilot screen of 26,000 small molecules to look for cell wall-active chemicals in real time using an autonomous luminescence gene cluster driven by the promoter of ywaC, which encodes a guanosine tetra(penta)phosphate synthetase that is expressed under cell wall stress. The promoter-reporter system was generally much more sensitive than growth inhibition testing and responded almost exclusively to cell wall-active antibiotics. Follow-up testing of the compounds from the pilot screen with secondary assays to verify the mechanism of action led to the discovery of 9 novel cell wall-active compounds. PMID:24687489

Modifying lignin to improve bioenergy feedstocks: strengthening the barrier against pathogens?

USDA-ARS?s Scientific Manuscript database

Lignin is a ubiquitous polymer present in cell walls of all vascular plants, where it rigidifies and strengthens the cell wall structure through covalent cross-linkages to cell wall polysaccharides. The presence of lignin makes the cell wall recalcitrant to conversion into fermentable sugars for bi...

A new Caenorhabditis elegans model of human huntingtin 513 aggregation and toxicity in body wall muscles.

PubMed

Lee, Amy L; Ung, Hailey M; Sands, L Paul; Kikis, Elise A

2017-01-01

Expanded polyglutamine repeats in different proteins are the known determinants of at least nine progressive neurodegenerative disorders whose symptoms include cognitive and motor impairment that worsen as patients age. One such disorder is Huntington's Disease (HD) that is caused by a polyglutamine expansion in the human huntingtin protein (htt). The polyglutamine expansion destabilizes htt leading to protein misfolding, which in turn triggers neurodegeneration and the disruption of energy metabolism in muscle cells. However, the molecular mechanisms that underlie htt proteotoxicity have been somewhat elusive, and the muscle phenotypes have not been well studied. To generate tools to elucidate the basis for muscle dysfunction, we engineered Caenorhabditis elegans to express a disease-associated 513 amino acid fragment of human htt in body wall muscle cells. We show that this htt fragment aggregates in C. elegans in a polyglutamine length-dependent manner and is toxic. Toxicity manifests as motor impairment and a shortened lifespan. Compared to previous models, the data suggest that the protein context in which a polyglutamine tract is embedded alters aggregation propensity and toxicity, likely by affecting interactions with the muscle cell environment.

Lower cell wall pectin solubilisation and galactose loss during early fruit development in apple (Malus x domestica) cultivar 'Scifresh' are associated with slower softening rate.

PubMed

Ng, Jovyn K T; Schröder, Roswitha; Brummell, David A; Sutherland, Paul W; Hallett, Ian C; Smith, Bronwen G; Melton, Laurence D; Johnston, Jason W

2015-03-15

Substantial differences in softening behaviour can exist between fruit even within the same species. Apple cultivars 'Royal Gala' and 'Scifresh' soften at different rates despite having a similar genetic background and producing similar amounts of ethylene during ripening. An examination of cell wall metabolism from the fruitlet to the ripe stages showed that in both cultivars pectin solubilisation increased during cell expansion, declined at the mature stage and then increased again during ripening. This process was much less pronounced in the slower softening 'Scifresh' than in 'Royal Gala' at every developmental stage examined, consistent with less cell separation and softening in this cultivar. Both cultivars also exhibited a progressive loss of pectic galactan and arabinan side chains during development. The cell wall content of arabinose residues was similar in both cultivars, but the galactose residue content in 'Scifresh' remained higher than that of 'Royal Gala' at every developmental stage. The higher content of cell wall galactose residue in 'Scifresh' cell walls correlated with a lower β-galactosidase activity and more intense immunolabelling of RG-I galactan side chains in both microscopy sections and glycan microarrays. A high cell wall galactan content has been associated with reduced cell wall porosity, which may restrict access of cell wall-modifying enzymes and thus maintain better structural integrity later in development. The data suggest that the composition and structure of the cell wall at very early development stages may influence subsequent cell wall loosening, and may even predispose the wall's ensuing properties. Copyright © 2014 Elsevier GmbH. All rights reserved.

Cell wall integrity modulates RHO1 activity via the exchange factor ROM2.

PubMed Central

Bickle, M; Delley, P A; Schmidt, A; Hall, M N

1998-01-01

The essential phosphatidylinositol kinase homologue TOR2 of Saccharomyces cerevisiae controls the actin cytoskeleton by activating a GTPase switch consisting of RHO1 (GTPase), ROM2 (GEF) and SAC7 (GAP). We have identified two mutations, rot1-1 and rot2-1, that suppress the loss of TOR2 and are synthetic-lethal. The wild-type ROT1 and ROT2 genes and a multicopy suppressor, BIG1, were isolated by their ability to rescue the rot1-1 rot2-1 double mutant. ROT2 encodes glucosidase II, and ROT1 and BIG1 encode novel proteins. We present evidence that cell wall defects activate RHO1. First, rot1, rot2, big1, cwh41, gas1 and fks1 mutations all confer cell wall defects and suppress tor2(ts). Second, destabilizing the cell wall by supplementing the growth medium with 0.005% SDS also suppresses a tor2(ts) mutation. Third, disturbing the cell wall with SDS or a rot1, rot2, big1, cwh41, gas1 or fks1 mutation increases GDP/GTP exchange activity toward RHO1. These results suggest that cell wall defects suppress a tor2 mutation by activating RHO1 independently of TOR2, thereby inducing TOR2-independent polarization of the actin cytoskeleton and cell wall synthesis. Activation of RHO1, a subunit of the cell wall synthesis enzyme glucan synthase, by a cell wall alteration would ensure that cell wall synthesis occurs only when and where needed. The mechanism of RHO1 activation by a cell wall alteration is via the exchange factor ROM2 and could be analogous to signalling by integrin receptors in mammalian cells. PMID:9545237

DISTRIBUTION OF RADIOACTIVITY IN AUTOLYZED CELL WALL OF BACILLUS CEREUS DURING SPHEROPLAST FORMATION1

PubMed Central

Kronish, Donald P.; Mohan, Raam R.; Schwartz, Benjamin S.

1964-01-01

Kronish, Donald P. (Warner-Lambert Research Institute, Morris Plains, N.J.), Raam R. Mohan, and Benjamin S. Schwartz. Distribution of radioactivity in autolyzed cell wall of Bacillus cereus during spheroplast formation. J. Bacteriol. 87:581–587. 1964.—Spheroplasts of Bacillus cereus strain T were produced from cells grown in the presence of uniformly labeled C14-glucose. At regular intervals during spheroplast formation, enzymatically degraded cell wall was isolated by a new procedure. Radioactivity of solubilized cell wall in cell-free material increased from 2.5 to 42% of the total incorporated label during spheroplast formation. The rate of cell-wall degradation as measured by increase in radioactivity was biphasic with relative slopes of 2.0 and 5.0. During autolytic depolymerization of B. cereus cell wall, two major components were solubilized at different rates. Chemical fractionation revealed these to be a peptide and a mucopeptide. The possibility of two enzymes being involved in spheroplast formation and cell-wall degradation is discussed. Images PMID:14127573

Suppression of Hydroxycinnamate Network Formation in Cell Walls of Rice Shoots Grown under Microgravity Conditions in Space

PubMed Central

Wakabayashi, Kazuyuki; Soga, Kouichi; Hoson, Takayuki; Kotake, Toshihisa; Yamazaki, Takashi; Higashibata, Akira; Ishioka, Noriaki; Shimazu, Toru; Fukui, Keiji; Osada, Ikuko; Kasahara, Haruo; Kamada, Motoshi

2015-01-01

Network structures created by hydroxycinnamate cross-links within the cell wall architecture of gramineous plants make the cell wall resistant to the gravitational force of the earth. In this study, the effects of microgravity on the formation of cell wall-bound hydroxycinnamates were examined using etiolated rice shoots simultaneously grown under artificial 1 g and microgravity conditions in the Cell Biology Experiment Facility on the International Space Station. Measurement of the mechanical properties of cell walls showed that shoot cell walls became stiff during the growth period and that microgravity suppressed this stiffening. Amounts of cell wall polysaccharides, cell wall-bound phenolic acids, and lignin in rice shoots increased as the shoot grew. Microgravity did not influence changes in the amounts of cell wall polysaccharides or phenolic acid monomers such as ferulic acid (FA) and p-coumaric acid, but it suppressed increases in diferulic acid (DFA) isomers and lignin. Activities of the enzymes phenylalanine ammonia-lyase (PAL) and cell wall-bound peroxidase (CW-PRX) in shoots also increased as the shoot grew. PAL activity in microgravity-grown shoots was almost comparable to that in artificial 1 g-grown shoots, while CW-PRX activity increased less in microgravity-grown shoots than in artificial 1 g-grown shoots. Furthermore, the increases in expression levels of some class III peroxidase genes were reduced under microgravity conditions. These results suggest that a microgravity environment modifies the expression levels of certain class III peroxidase genes in rice shoots, that the resultant reduction of CW-PRX activity may be involved in suppressing DFA formation and lignin polymerization, and that this suppression may cause a decrease in cross-linkages within the cell wall architecture. The reduction in intra-network structures may contribute to keeping the cell wall loose under microgravity conditions. PMID:26378793

Estimation of the physiological mechanical conditioning in vascular tissue engineering by a predictive fluid-structure interaction approach.

PubMed

Tresoldi, Claudia; Bianchi, Elena; Pellegata, Alessandro Filippo; Dubini, Gabriele; Mantero, Sara

2017-08-01

The in vitro replication of physiological mechanical conditioning through bioreactors plays a crucial role in the development of functional Small-Caliber Tissue-Engineered Blood Vessels. An in silico scaffold-specific model under pulsatile perfusion provided by a bioreactor was implemented using a fluid-structure interaction (FSI) approach for viscoelastic tubular scaffolds (e.g. decellularized swine arteries, DSA). Results of working pressures, circumferential deformations, and wall shear stress on DSA fell within the desired physiological range and indicated the ability of this model to correctly predict the mechanical conditioning acting on the cells-scaffold system. Consequently, the FSI model allowed us to a priori define the stimulation pattern, driving in vitro physiological maturation of scaffolds, especially with viscoelastic properties.

Energy biotechnology in the CRISPR-Cas9 era.

PubMed

Estrela, Raissa; Cate, Jamie Harrison Doudna

2016-04-01

The production of bioenergy from plant biomass previously relied on using microorganisms that rapidly and efficiently convert simple sugars into fuels and chemicals. However, to exploit the far more abundant carbon fixed in plant cell walls, future industrial production hosts will need to be engineered to leverage the most efficient biochemical pathways and most robust traits that can be found in nature. The CRISPR-Cas9 genome editing technology now enables writing the genome at will, which will allow biotechnology to become an 'information science.' This review covers recent advances in using CRISPR-Cas9 to engineer the genomes of a wide variety of organisms that could be use in the industrial production of biofuels and renewable chemicals. Copyright © 2016 Elsevier Ltd. All rights reserved.

Microanalysis of plant cell wall polysaccharides.

PubMed

Obel, Nicolai; Erben, Veronika; Schwarz, Tatjana; Kühnel, Stefan; Fodor, Andrea; Pauly, Markus

2009-09-01

Oligosaccharide Mass Profiling (OLIMP) allows a fast and sensitive assessment of cell wall polymer structure when coupled with Matrix Assisted Laser Desorption Ionisation Time Of Flight Mass Spectrometry (MALDI-TOF MS). The short time required for sample preparation and analysis makes possible the study of a wide range of plant organs, revealing a high degree of heterogeneity in the substitution pattern of wall polymers such as the cross-linking glycan xyloglucan and the pectic polysaccharide homogalacturonan. The high sensitivity of MALDI-TOF allows the use of small amounts of samples, thus making it possible to investigate the wall structure of single cell types when material is collected by such methods as laser micro-dissection. As an example, the analysis of the xyloglucan structure in the leaf cell types outer epidermis layer, entire epidermis cell layer, palisade mesophyll cells, and vascular bundles were investigated. OLIMP is amenable to in situ wall analysis, where wall polymers are analyzed on unprepared plant tissue itself without first isolating cell walls. In addition, OLIMP enables analysis of wall polymers in Golgi-enriched fractions, the location of nascent matrix polysaccharide biosynthesis, enabling separation of the processes of wall biosynthesis versus post-deposition apoplastic metabolism. These new tools will make possible a semi-quantitative analysis of the cell wall at an unprecedented level.

Genetic engineering of woody plants: current and future targets in a stressful environment.

PubMed

Osakabe, Yuriko; Kajita, Shinya; Osakabe, Keishi

2011-06-01

Abiotic stress is a major factor in limiting plant growth and productivity. Environmental degradation, such as drought and salinity stresses, will become more severe and widespread in the world. To overcome severe environmental stress, plant biotechnologies, such as genetic engineering in woody plants, need to be implemented. The adaptation of plants to environmental stress is controlled by cascades of molecular networks including cross-talk with other stress signaling mechanisms. The present review focuses on recent studies concerning genetic engineering in woody plants for the improvement of the abiotic stress responses. Furthermore, it highlights the recent advances in the understanding of molecular responses to stress. The review also summarizes the basis of a molecular mechanism for cell wall biosynthesis and the plant hormone responses to regulate tree growth and biomass in woody plants. This would facilitate better understanding of the control programs of biomass production under stressful conditions. Copyright © Physiologia Plantarum 2011.

The fabrication of PLGA microvessel scaffolds with nano-patterned inner walls.

PubMed

Wang, Gou-Jen; Lin, Yan-Cheng; Hsu, Shan-Hui

2010-10-01

Poly (lactic-co-glycolic acid) (PLGA) is one of the most commonly used biodegradable, biocompatible materials. Nanostructured PLGA has immense potential for application in tissue engineering. In this article we discuss a novel approach for the fabrication of PLGA microvessel scaffolds with nanostructured inner walls. In this novel nano-patterning approach, the thermal reflow technique is first adapted to fabricate a semi-cylindrical photoresist master mold. A thin film of titanium and a thin film of aluminum are sputtered in sequence on the semi-cylindrical microvessel network. Aluminum foil anodization is then executed to transform the aluminum thin film into a porous anodic aluminum oxide (AAO) film. During the casting process a PLGA solution is cast on the AAO film to build up semi-cylindrical PLGA microstructures with nanostructured inner walls after which inductive coupled plasma (ICP) is implemented to assist bonding of the two PLGA structures. The result is the building of a network of microchannels with nano-patterned inner walls. Bovine endothelial cells (BECs) are carefully cultured in the scaffold via semi-dynamic seeding for 7 days. Observations show that the BECs grew more separately in a nano-patterned microvessel scaffold than they did in a smooth surface scaffold.

Short-Term Boron Deprivation Inhibits Endocytosis of Cell Wall Pectins in Meristematic Cells of Maize and Wheat Root Apices1

PubMed Central

Yu, Qin; Hlavacka, Andrej; Matoh, Toru; Volkmann, Dieter; Menzel, Diedrik; Goldbach, Heiner E.; Baluška, František

2002-01-01

By using immunofluorescence microscopy, we observed rapidly altered distribution patterns of cell wall pectins in meristematic cells of maize (Zea mays) and wheat (Triticum aestivum) root apices. This response was shown for homogalacturonan pectins characterized by a low level (up to 40%) of methylesterification and for rhamnogalacturonan II pectins cross-linked by a borate diol diester. Under boron deprivation, abundance of these pectins rapidly increased in cell walls, whereas their internalization was inhibited, as evidenced by a reduced and even blocked accumulation of these cell wall pectins within brefeldin A-induced compartments. In contrast, root cells of species sensitive to the boron deprivation, like zucchini (Cucurbita pepo) and alfalfa (Medicago sativa), do not internalize cell wall pectins into brefeldin A compartments and do not show accumulation of pectins in their cell walls under boron deprivation. For maize and wheat root apices, we favor an apoplastic target for the primary action of boron deprivation, which signals deeper into the cell via endocytosis-mediated pectin signaling along putative cell wall-plasma membrane-cytoskeleton continuum. PMID:12226520

Fish cell lines as a tool for the ecotoxicity assessment and ranking of engineered nanomaterials.

PubMed

Bermejo-Nogales, A; Fernández-Cruz, M L; Navas, J M

2017-11-01

Risk assessment of engineered nanomaterials (ENMs) is being hindered by the sheer production volume of these materials. In this regard, the grouping and ranking of ENMs appears as a promising strategy. Here we sought to evaluate the usefulness of in vitro systems based on fish cell lines for ranking a set of ENMs on the basis of their cytotoxicity. We used the topminnow (Poeciliopsis lucida) liver cell line (PLHC-1) and the rainbow trout (Oncorhynchus mykiss) fibroblast-like gonadal cell line (RTG-2). ENMs were obtained from the EU Joint Research Centre repository. The size frequency distribution of ENM suspensions in cell culture media was characterized. Cytotoxicity was evaluated after 24 h of exposure. PLHC-1 cells exhibited higher sensitivity to the ENMs than RTG-2 cells. ZnO-NM was found to exert toxicity mainly by altering lysosome function and metabolic activity, while multi-walled carbon nanotubes (MWCNTs) caused plasma membrane disruption at high concentrations. The hazard ranking for toxicity (ZnO-NM > MWCNT ≥ CeO 2 -NM = SiO 2 -NM) was inversely related to the ranking in size detected in culture medium. Our findings reveal the suitability of fish cell lines for establishing hazard rankings of ENMs in the framework of integrated approaches to testing and assessment. Copyright © 2017 Elsevier Inc. All rights reserved.

Cell wall composition profiling of parasitic giant dodder (Cuscuta reflexa) and its hosts: a priori differences and induced changes.

PubMed

Johnsen, Hanne R; Striberny, Bernd; Olsen, Stian; Vidal-Melgosa, Silvia; Fangel, Jonatan U; Willats, William G T; Rose, Jocelyn K C; Krause, Kirsten

2015-08-01

Host plant penetration is the gateway to survival for holoparasitic Cuscuta and requires host cell wall degradation. Compositional differences of cell walls may explain why some hosts are amenable to such degradation while others can resist infection. Antibody-based techniques for comprehensive profiling of cell wall epitopes and cell wall-modifying enzymes were applied to several susceptible hosts and a resistant host of Cuscuta reflexa and to the parasite itself. Infected tissue of Pelargonium zonale contained high concentrations of de-esterified homogalacturonans in the cell walls, particularly adjacent to the parasite's haustoria. High pectinolytic activity in haustorial extracts and high expression levels of pectate lyase genes suggest that the parasite contributes directly to wall remodeling. Mannan and xylan concentrations were low in P. zonale and in five susceptible tomato introgression lines, but high in the resistant Solanum lycopersicum cv M82, and in C. reflexa itself. Knowledge of the composition of resistant host cell walls and the parasite's own cell walls is useful in developing strategies to prevent infection by parasitic plants. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

The chaotrope-soluble glycoprotein GP1 is a constituent of the insoluble glycoprotein framework of the Chlamydomonas cell wall.

PubMed

Voigt, Jürgen; Frank, Ronald; Wöstemeyer, Johannes

2009-02-01

Chlamydomonas reinhardtii wild-type cells are surrounded by the insoluble cell wall component, a sac-like framework of cross-linked glycoproteins containing 22% hydroxyproline. The chaotrope-soluble cell wall glycoprotein GP1 is the only polypeptide with an even higher proportion of hydroxyproline (35%) occurring in vegetative C. reinhardtii cells. Mass spectrometric analyses of peptides released from the purified insoluble cell wall fraction by trypsin treatment and epitope analyses of polyclonal antibodies raised against different deglycosylation products of this particular wall fraction using 181 chemically synthesized GP1-derived pentadecapeptides revealed evidence that GP1 is indeed a constituent of the insoluble wall component.

Calcium deprivation disrupts enlargement of Chara corallina cells: further evidence for the calcium pectate cycle.

PubMed

Proseus, Timothy E; Boyer, John S

2012-06-01

Pectin is a normal constituent of cell walls of green plants. When supplied externally to live cells or walls isolated from the large-celled green alga Chara corallina, pectin removes calcium from load-bearing cross-links in the wall, loosening the structure and allowing it to deform more rapidly under the action of turgor pressure. New Ca(2+) enters the vacated positions in the wall and the externally supplied pectin binds to the wall, depositing new wall material that strengthens the wall. A calcium pectate cycle has been proposed for these sub-reactions. In the present work, the cycle was tested in C. corallina by depriving the wall of external Ca(2+) while allowing the cycle to run. The prediction is that growth would eventually be disrupted by a lack of adequate deposition of new wall. The test involved adding pectate or the calcium chelator EGTA to the Ca(2+)-containing culture medium to bind the calcium while the cycle ran in live cells. After growth accelerated, turgor and growth eventually decreased, followed by an abrupt turgor loss and growth cessation. The same experiment with isolated walls suggested the walls of live cells became unable to support the plasma membrane. If instead the pectate or EGTA was replaced with fresh Ca(2+)-containing culture medium during the initial acceleration in live cells, growth was not disrupted and returned to the original rates. The operation of the cycle was thus confirmed, providing further evidence that growth rates and wall biosynthesis are controlled by these sub-reactions in plant cell walls.

Branched Pectic Galactan in Phloem-Sieve-Element Cell Walls: Implications for Cell Mechanics.

PubMed

Torode, Thomas A; O'Neill, Rachel; Marcus, Susan E; Cornuault, Valérie; Pose, Sara; Lauder, Rebecca P; Kračun, Stjepan K; Rydahl, Maja Gro; Andersen, Mathias C F; Willats, William G T; Braybrook, Siobhan A; Townsend, Belinda J; Clausen, Mads H; Knox, J Paul

2018-02-01

A major question in plant biology concerns the specification and functional differentiation of cell types. This is in the context of constraints imposed by networks of cell walls that both adhere cells and contribute to the form and function of developing organs. Here, we report the identification of a glycan epitope that is specific to phloem sieve element cell walls in several systems. A monoclonal antibody, designated LM26, binds to the cell wall of phloem sieve elements in stems of Arabidopsis ( Arabidopsis thaliana ), Miscanthus x giganteus , and notably sugar beet ( Beta vulgaris ) roots where phloem identification is an important factor for the study of phloem unloading of Suc. Using microarrays of synthetic oligosaccharides, the LM26 epitope has been identified as a β-1,6-galactosyl substitution of β-1,4-galactan requiring more than three backbone residues for optimized recognition. This branched galactan structure has previously been identified in garlic ( Allium sativum ) bulbs in which the LM26 epitope is widespread throughout most cell walls including those of phloem cells. Garlic bulb cell wall material has been used to confirm the association of the LM26 epitope with cell wall pectic rhamnogalacturonan-I polysaccharides. In the phloem tissues of grass stems, the LM26 epitope has a complementary pattern to that of the LM5 linear β-1,4-galactan epitope, which is detected only in companion cell walls. Mechanical probing of transverse sections of M x giganteus stems and leaves by atomic force microscopy indicates that phloem sieve element cell walls have a lower indentation modulus (indicative of higher elasticity) than companion cell walls. © 2018 The author(s). All Rights Reserved.

Viscoelastic properties of cell walls of single living plant cells determined by dynamic nanoindentation

PubMed Central

Hayot, Céline M.; Forouzesh, Elham; Goel, Ashwani; Avramova, Zoya; Turner, Joseph A.

2012-01-01

Plant development results from controlled cell divisions, structural modifications, and reorganizations of the cell wall. Thereby, regulation of cell wall behaviour takes place at multiple length scales involving compositional and architectural aspects in addition to various developmental and/or environmental factors. The physical properties of the primary wall are largely determined by the nature of the complex polymer network, which exhibits time-dependent behaviour representative of viscoelastic materials. Here, a dynamic nanoindentation technique is used to measure the time-dependent response and the viscoelastic behaviour of the cell wall in single living cells at a micron or sub-micron scale. With this approach, significant changes in storage (stiffness) and loss (loss of energy) moduli are captured among the tested cells. The results reveal hitherto unknown differences in the viscoelastic parameters of the walls of same-age similarly positioned cells of the Arabidopsis ecotypes (Col 0 and Ws 2). The technique is also shown to be sensitive enough to detect changes in cell wall properties in cells deficient in the activity of the chromatin modifier ATX1. Extensive computational modelling of the experimental measurements (i.e. modelling the cell as a viscoelastic pressure vessel) is used to analyse the influence of the wall thickness, as well as the turgor pressure, at the positions of our measurements. By combining the nanoDMA technique with finite element simulations quantifiable measurements of the viscoelastic properties of plant cell walls are achieved. Such techniques are expected to find broader applications in quantifying the influence of genetic, biological, and environmental factors on the nanoscale mechanical properties of the cell wall. PMID:22291130

Reconstitution of a secondary cell wall in a secondary cell wall-deficient Arabidopsis mutant.

PubMed

Sakamoto, Shingo; Mitsuda, Nobutaka

2015-02-01

The secondary cell wall constitutes a rigid frame of cells in plant tissues where rigidity is required. Deposition of the secondary cell wall in fiber cells contributes to the production of wood in woody plants. The secondary cell wall is assembled through co-operative activities of many enzymes, and their gene expression is precisely regulated by a pyramidal cascade of transcription factors. Deposition of a transmuted secondary cell wall in empty fiber cells by expressing selected gene(s) in this cascade has not been attempted previously. In this proof-of-concept study, we expressed chimeric activators of 24 transcription factors that are preferentially expressed in the stem, in empty fiber cells of the Arabidopsis nst1-1 nst3-1 double mutant, which lacks a secondary cell wall in fiber cells, under the control of the NST3 promoter. The chimeric activators of MYB46, SND2 and ANAC075, as well as NST3, reconstituted a secondary cell wall with different characteristics from those of the wild type in terms of its composition. The transgenic lines expressing the SND2 or ANAC075 chimeric activator showed increased glucose and xylose, and lower lignin content, whereas the transgenic line expressing the MYB46 chimeric activator showed increased mannose content. The expression profile of downstream genes in each transgenic line was also different from that of the wild type. This study proposed a new screening strategy to identify factors of secondary wall formation and also suggested the potential of the artificially reconstituted secondary cell walls as a novel raw material for production of bioethanol and other chemicals. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

Changes in levels of cell wall constituents in wheat seedlings grown under continuous hypergravity conditions

NASA Astrophysics Data System (ADS)

Wakabayashi, K.; Soga, K.; Kamisaka, S.; Hoson, T.

Effects of continuous hypergravity stimuli on the amounts and composition of cell wall constituents were investigated in wheat shoots. Hypergravity (300 g) treatment for three days after germination increased the net amount of cell wall polysaccharides such as hemicellulose and cellulose, but reduced the shoot elongation. As a result, the amount of cell wall polysaccharides per unit length of shoot increased under hypergravity. The hemicellulose fraction contained polysaccharides in the middle and low molecular mass range (5 kDa-1 MDa) and increased in response to hypergravity. Also, the amounts of arabinose (Ara) and xylose (Xyl), the major sugar components of the hemicellulose fraction, increased under hypergravity conditions. In addition to wall polysaccharides, hypergravity increased the amounts of cell wall-bound phenolic acids, such as ferulic acid (FA) and diferulic acid (DFA). Furthermore, the activity of phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) was enhanced under hypergravity conditions. These results suggest that continuous hypergravity stimulates the synthesis of cell wall constituents, especially hemicellulosic arabinoxylans and cell wall-bound FA and DFA in wheat shoots. The increased PAL activity may promote the formation of FA and DFA. These changes in cell wall architecture may be involved in making rigid and tough cell walls under hypergravity conditions and thereby contribute to the ability of plant to sustain their structures against gravitational stimuli.

Electron Tomography of Cryo-Immobilized Plant Tissue: A Novel Approach to Studying 3D Macromolecular Architecture of Mature Plant Cell Walls In Situ

PubMed Central

Sarkar, Purbasha; Bosneaga, Elena; Yap, Edgar G.; Das, Jyotirmoy; Tsai, Wen-Ting; Cabal, Angelo; Neuhaus, Erica; Maji, Dolonchampa; Kumar, Shailabh; Joo, Michael; Yakovlev, Sergey; Csencsits, Roseann; Yu, Zeyun; Bajaj, Chandrajit; Downing, Kenneth H.; Auer, Manfred

2014-01-01

Cost-effective production of lignocellulosic biofuel requires efficient breakdown of cell walls present in plant biomass to retrieve the wall polysaccharides for fermentation. In-depth knowledge of plant cell wall composition is therefore essential for improving the fuel production process. The precise spatial three-dimensional (3D) organization of cellulose, hemicellulose, pectin and lignin within plant cell walls remains unclear to date since the microscopy techniques used so far have been limited to two-dimensional, topographic or low-resolution imaging, or required isolation or chemical extraction of the cell walls. In this paper we demonstrate that by cryo-immobilizing fresh tissue, then either cryo-sectioning or freeze-substituting and resin embedding, followed by cryo- or room temperature (RT) electron tomography, respectively, we can visualize previously unseen details of plant cell wall architecture in 3D, at macromolecular resolution (∼2 nm), and in near-native state. Qualitative and quantitative analyses showed that wall organization of cryo-immobilized samples were preserved remarkably better than conventionally prepared samples that suffer substantial extraction. Lignin-less primary cell walls were well preserved in both self-pressurized rapidly frozen (SPRF), cryo-sectioned samples as well as high-pressure frozen, freeze-substituted and resin embedded (HPF-FS-resin) samples. Lignin-rich secondary cell walls appeared featureless in HPF-FS-resin sections presumably due to poor stain penetration, but their macromolecular features could be visualized in unprecedented details in our cryo-sections. While cryo-tomography of vitreous tissue sections is currently proving to be instrumental in developing 3D models of lignin-rich secondary cell walls, here we confirm that the technically easier method of RT-tomography of HPF-FS-resin sections could be used immediately for routine study of low-lignin cell walls. As a proof of principle, we characterized the primary cell walls of a mutant (cob-6) and wild type Arabidopsis hypocotyl parenchyma cells by RT-tomography of HPF-FS-resin sections, and detected a small but significant difference in spatial organization of cellulose microfibrils in the mutant walls. PMID:25207917

Tools to Understand Structural Property Relationships for Wood Cell Walls

Treesearch

Joseph E. Jakes; Daniel J. Yelle; Charles R. Frihart

2011-01-01

Understanding structure-property relationships for wood cell walls has been hindered by the complex polymeric structures comprising these cell walls and the difficulty in assessing meaningful mechanical property measurements of individual cell walls. To help overcome these hindrances, we have developed two experimental methods: 1) two-dimensional solution state nuclear...

The plant cell wall integrity maintenance mechanism--a case study of a cell wall plasma membrane signaling network.

PubMed

Hamann, Thorsten

2015-04-01

Some of the most important functions of plant cell walls are protection against biotic/abiotic stress and structural support during growth and development. A prerequisite for plant cell walls to perform these functions is the ability to perceive different types of stimuli in both qualitative and quantitative manners and initiate appropriate responses. The responses in turn involve adaptive changes in cellular and cell wall metabolism leading to modifications in the structures originally required for perception. While our knowledge about the underlying plant mechanisms is limited, results from Saccharomyces cerevisiae suggest the cell wall integrity maintenance mechanism represents an excellent example to illustrate how the molecular mechanisms responsible for stimulus perception, signal transduction and integration can function. Here I will review the available knowledge about the yeast cell wall integrity maintenance system for illustration purposes, summarize the limited knowledge available about the corresponding plant mechanism and discuss the relevance of the plant cell wall integrity maintenance mechanism in biotic stress responses. Copyright © 2014 Elsevier Ltd. All rights reserved.

Building and degradation of secondary cell walls: are there common patterns of lamellar assembly of cellulose microfibrils and cell wall delamination?

PubMed

De Micco, Veronica; Ruel, Katia; Joseleau, Jean-Paul; Aronne, Giovanna

2010-08-01

During cell wall formation and degradation, it is possible to detect cellulose microfibrils assembled into thicker and thinner lamellar structures, respectively, following inverse parallel patterns. The aim of this study was to analyse such patterns of microfibril aggregation and cell wall delamination. The thickness of microfibrils and lamellae was measured on digital images of both growing and degrading cell walls viewed by means of transmission electron microscopy. To objectively detect, measure and classify microfibrils and lamellae into thickness classes, a method based on the application of computerized image analysis combined with graphical and statistical methods was developed. The method allowed common classes of microfibrils and lamellae in cell walls to be identified from different origins. During both the formation and degradation of cell walls, a preferential formation of structures with specific thickness was evidenced. The results obtained with the developed method allowed objective analysis of patterns of microfibril aggregation and evidenced a trend of doubling/halving lamellar structures, during cell wall formation/degradation in materials from different origin and which have undergone different treatments.

Cell wall-bound silicon optimizes ammonium uptake and metabolism in rice cells.

PubMed

Sheng, Huachun; Ma, Jie; Pu, Junbao; Wang, Lijun

2018-05-16

Turgor-driven plant cell growth depends on cell wall structure and mechanics. Strengthening of cell walls on the basis of an association and interaction with silicon (Si) could lead to improved nutrient uptake and optimized growth and metabolism in rice (Oryza sativa). However, the structural basis and physiological mechanisms of nutrient uptake and metabolism optimization under Si assistance remain obscure. Single-cell level biophysical measurements, including in situ non-invasive micro-testing (NMT) of NH4+ ion fluxes, atomic force microscopy (AFM) of cell walls, and electrolyte leakage and membrane potential, as well as whole-cell proteomics using isobaric tags for relative and absolute quantification (iTRAQ), were performed. The altered cell wall structure increases the uptake rate of the main nutrient NH4+ in Si-accumulating cells, whereas the rate is only half in Si-deprived counterparts. Rigid cell walls enhanced by a wall-bound form of Si as the structural basis stabilize cell membranes. This, in turn, optimizes nutrient uptake of the cells in the same growth phase without any requirement for up-regulation of transmembrane ammonium transporters. Optimization of cellular nutrient acquisition strategies can substantially improve performance in terms of growth, metabolism and stress resistance.

The Draft Genome of the Invasive Walking Stick, Medauroidea extradendata, Reveals Extensive Lineage-Specific Gene Family Expansions of Cell Wall Degrading Enzymes in Phasmatodea

PubMed Central

Brand, Philipp; Lin, Wei; Johnson, Brian R.

2018-01-01

Plant cell wall components are the most abundant macromolecules on Earth. The study of the breakdown of these molecules is thus a central question in biology. Surprisingly, plant cell wall breakdown by herbivores is relatively poorly understood, as nearly all early work focused on the mechanisms used by symbiotic microbes to breakdown plant cell walls in insects such as termites. Recently, however, it has been shown that many organisms make endogenous cellulases. Insects, and other arthropods, in particular have been shown to express a variety of plant cell wall degrading enzymes in many gene families with the ability to break down all the major components of the plant cell wall. Here we report the genome of a walking stick, Medauroidea extradentata, an obligate herbivore that makes uses of endogenously produced plant cell wall degrading enzymes. We present a draft of the 3.3Gbp genome along with an official gene set that contains a diversity of plant cell wall degrading enzymes. We show that at least one of the major families of plant cell wall degrading enzymes, the pectinases, have undergone a striking lineage-specific gene family expansion in the Phasmatodea. This genome will be a useful resource for comparative evolutionary studies with herbivores in many other clades and will help elucidate the mechanisms by which metazoans breakdown plant cell wall components. PMID:29588379

Plant cell walls throughout evolution: towards a molecular understanding of their design principles.

PubMed

Sarkar, Purbasha; Bosneaga, Elena; Auer, Manfred

2009-01-01

Throughout their life, plants typically remain in one location utilizing sunlight for the synthesis of carbohydrates, which serve as their sole source of energy as well as building blocks of a protective extracellular matrix, called the cell wall. During the course of evolution, plants have repeatedly adapted to their respective niche, which is reflected in the changes of their body plan and the specific design of cell walls. Cell walls not only changed throughout evolution but also are constantly remodelled and reconstructed during the development of an individual plant, and in response to environmental stress or pathogen attacks. Carbohydrate-rich cell walls display complex designs, which together with the presence of phenolic polymers constitutes a barrier for microbes, fungi, and animals. Throughout evolution microbes have co-evolved strategies for efficient breakdown of cell walls. Our current understanding of cell walls and their evolutionary changes are limited as our knowledge is mainly derived from biochemical and genetic studies, complemented by a few targeted yet very informative imaging studies. Comprehensive plant cell wall models will aid in the re-design of plant cell walls for the purpose of commercially viable lignocellulosic biofuel production as well as for the timber, textile, and paper industries. Such knowledge will also be of great interest in the context of agriculture and to plant biologists in general. It is expected that detailed plant cell wall models will require integrated correlative multimodal, multiscale imaging and modelling approaches, which are currently underway.

Differential growth of pavement cells of Arabidopsis thaliana leaf epidermis as revealed by microbead labeling.

PubMed

Elsner, Joanna; Lipowczan, Marcin; Kwiatkowska, Dorota

2018-02-01

In numerous vascular plants, pavement cells of the leaf epidermis are shaped like a jigsaw-puzzle piece. Knowledge about the subcellular pattern of growth that accompanies morphogenesis of such a complex shape is crucial for studies of the role of the cytoskeleton, cell wall and phytohormones in plant cell development. Because the detailed growth pattern of the anticlinal and periclinal cell walls remains unknown, our aim was to measure pavement cell growth at a subcellular resolution. Using fluorescent microbeads applied to the surface of the adaxial leaf epidermis of Arabidopsis thaliana as landmarks for growth computation, we directly assessed the growth rates for the outer periclinal and anticlinal cell walls at a subcellular scale. We observed complementary tendencies in the growth pattern of the outer periclinal and anticlinal cell walls. Central portions of periclinal walls were characterized by relatively slow growth, while growth of the other wall portions was heterogeneous. Local growth of the periclinal walls accompanying lobe development after initiation was relatively fast and anisotropic, with maximal extension usually in the direction along the lobe axis. This growth pattern of the periclinal walls was complemented by the extension of the anticlinal walls, which was faster on the lobe sides than at the tips. Growth of the anticlinal and outer periclinal walls of leaf pavement cells is heterogeneous. The growth of the lobes resembles cell elongation via diffuse growth rather than tip growth. © 2018 Botanical Society of America.

CFD modelling wall heat transfer inside a combustion chamber using ANSYS forte

NASA Astrophysics Data System (ADS)

Plengsa-ard, C.; Kaewbumrung, M.

2018-01-01

A computational model has been performed to analyze a wall heat transfer in a single cylinder, direct injection and four-stroke diesel engine. A direct integration using detailed chemistry CHEMKIN is employed in a combustion model and the Reynolds Averaged Navier Stokes (RANS) turbulence model is used to simulate the flow in the cylinder. To obtain heat flux results, a modified classical variable-density wall heat transfer model is also performed. The model is validated using experimental data from a CUMMINs engine operated with a conventional diesel combustion. One operating engine condition is simulated. Comparisons of simulated in-cylinder pressure and heat release rates with experimental data shows that the model predicts the cylinder pressure and heat release rates reasonably well. The contour plot of instantaneous temperature are presented. Also, the contours of predicted heat flux results are shown. The magnitude of peak heat fluxes as predicted by the wall heat transfer model is in the range of the typical measure values in diesel combustion.

Structural characterization of a mixed-linkage glucan deficient mutant reveals alteration in cellulose microfibril orientation in rice coleoptile mesophyll cell walls

PubMed Central

Smith-Moritz, Andreia M.; Hao, Zhao; Fernández-Niño, Susana G.; Fangel, Jonatan U.; Verhertbruggen, Yves; Holman, Hoi-Ying N.; Willats, William G. T.; Ronald, Pamela C.; Scheller, Henrik V.; Heazlewood, Joshua L.; Vega-Sánchez, Miguel E.

2015-01-01

The CELLULOSE SYNTHASE-LIKE F6 (CslF6) gene was previously shown to mediate the biosynthesis of mixed-linkage glucan (MLG), a cell wall polysaccharide that is hypothesized to be tightly associated with cellulose and also have a role in cell expansion in the primary cell wall of young seedlings in grass species. We have recently shown that loss-of-function cslf6 rice mutants do not accumulate MLG in most vegetative tissues. Despite the absence of a structurally important polymer, MLG, these mutants are unexpectedly viable and only show a moderate growth compromise compared to wild type. Therefore these mutants are ideal biological systems to test the current grass cell wall model. In order to gain a better understanding of the role of MLG in the primary wall, we performed in-depth compositional and structural analyses of the cell walls of 3 day-old rice seedlings using various biochemical and novel microspectroscopic approaches. We found that cellulose content as well as matrix polysaccharide composition was not significantly altered in the MLG deficient mutant. However, we observed a significant change in cellulose microfibril bundle organization in mesophyll cell walls of the cslf6 mutant. Using synchrotron source Fourier Transform Mid-Infrared (FTM-IR) Spectromicroscopy for high-resolution imaging, we determined that the bonds associated with cellulose and arabinoxylan, another major component of the primary cell walls of grasses, were in a lower energy configuration compared to wild type, suggesting a slightly weaker primary wall in MLG deficient mesophyll cells. Taken together, these results suggest that MLG may influence cellulose deposition in mesophyll cell walls without significantly affecting anisotropic growth thus challenging MLG importance in cell wall expansion. PMID:26347754

Temperature modulates the cell wall mechanical properties of rice coleoptiles by altering the molecular mass of hemicellulosic polysaccharides

NASA Technical Reports Server (NTRS)

Nakamura, Yukiko; Wakabayashi, Kazuyuki; Hoson, Takayuki

2003-01-01

The present study was conducted to investigate the mechanism inducing the difference in the cell wall extensibility of rice (Oryza sativa L. cv. Koshihikari) coleoptiles grown under various temperature (10-50 degrees C) conditions. The growth rate and the cell wall extensibility of rice coleoptiles exhibited the maximum value at 30-40 degrees C, and became smaller as the growth temperature rose or dropped from this temperature range. The amounts of cell wall polysaccharides per unit length of coleoptile increased in coleoptiles grown at 40 degrees C, but not at other temperature conditions. On the other hand, the molecular size of hemicellulosic polysaccharides was small at temperatures where the cell wall extensibility was high (30-40 degrees C). The autolytic activities of cell walls obtained from coleoptiles grown at 30 and 40 degrees C were substantially higher than those grown at 10, 20 and 50 degrees C. Furthermore, the activities of (1-->3),(1-->4)-beta-glucanases extracted from coleoptile cell walls showed a similar tendency. When oat (1-->3),(1-->4)-beta-glucans with high molecular mass were incubated with the cell wall enzyme preparations from coleoptiles grown at various temperature conditions, the extensive molecular mass downshifts were brought about only by the cell wall enzymes obtained from coleoptiles grown at 30-40 degrees C. There were close correlations between the cell wall extensibility and the molecular mass of hemicellulosic polysaccharides or the activity of beta -glucanases. These results suggest that the environmental temperature regulates the cell wall extensibility of rice coleoptiles by modifying mainly the molecular mass of hemicellulosic polysaccharides. Modulation of the activity of beta-glucanases under various temperature conditions may be involved in the alteration of the molecular size of hemicellulosic polysaccharides.

The Complex Cell Wall Composition of Syncytia Induced by Plant Parasitic Cyst Nematodes Reflects Both Function and Host Plant.

PubMed

Zhang, Li; Lilley, Catherine J; Imren, Mustafa; Knox, J Paul; Urwin, Peter E

2017-01-01

Plant-parasitic cyst nematodes induce the formation of specialized feeding structures, syncytia, within their host roots. These unique plant organs serve as the sole nutrient resource for development and reproduction throughout the biotrophic interaction. The multinucleate syncytium, which arises through local dissolution of cell walls and protoplast fusion of multiple adjacent cells, has dense cytoplasm containing numerous organelles, surrounded by thickened outer cell walls that must withstand high turgor pressure. However, little is known about how the constituents of the syncytial cell wall and their conformation support its role during nematode parasitism. We used a set of monoclonal antibodies, targeted to a range of plant cell wall components, to reveal the microstructures of syncytial cell walls induced by four of the most economically important cyst nematode species, Globodera pallida , Heterodera glycines , Heterodera avenae and Heterodera filipjevi , in their respective potato, soybean, and spring wheat host roots. In situ fluorescence analysis revealed highly similar cell wall composition of syncytia induced by G. pallida and H. glycines . Both consisted of abundant xyloglucan, methyl-esterified homogalacturonan and pectic arabinan. In contrast, the walls of syncytia induced in wheat roots by H. avenae and H. filipjevi contain little xyloglucan but are rich in feruloylated xylan and arabinan residues, with variable levels of mixed-linkage glucan. The overall chemical composition of syncytial cell walls reflected the general features of root cell walls of the different host plants. We relate specific components of syncytial cell walls, such as abundant arabinan, methyl-esterification status of pectic homogalacturonan and feruloylation of xylan, to their potential roles in forming a network to support both the strength and flexibility required for syncytium function.

Structural characterization of a mixed-linkage glucan deficient mutant reveals alteration in cellulose microfibril orientation in rice coleoptile mesophyll cell walls

DOE PAGES

Smith-Moritz, Andreia M.; Hao, Zhao; Fernández-Nino, Susana G.; ...

2015-08-18

The CELLULOSE SYNTHASE-LIKE F6 (CslF6) gene was previously shown to mediate the biosynthesis of mixed-linkage glucan (MLG), a cell wall polysaccharide that is hypothesized to be tightly associated with cellulose and also have a role in cell expansion in the primary cell wall of young seedlings in grass species. We have recently shown that loss-of-function cslf6 rice mutants do not accumulate MLG in most vegetative tissues. Despite the absence of a structurally important polymer, MLG, these mutants are unexpectedly viable and only show a moderate growth compromise compared to wild type. Therefore these mutants are ideal biological systems to testmore » the current grass cell wall model. In order to gain a better understanding of the role of MLG in the primary wall, we performed in-depth compositional and structural analyses of the cell walls of 3 day-old rice seedlings using various biochemical and novel microspectroscopic approaches. We found that cellulose content as well as matrix polysaccharide composition was not significantly altered in the MLG deficient mutant. However, we observed a significant change in cellulose microfibril bundle organization in mesophyll cell walls of the cslf6 mutant. Using synchrotron source Fourier Transform Mid-Infrared (FTM-IR) Spectromicroscopy for high-resolution imaging, we determined that the bonds associated with cellulose and arabinoxylan, another major component of the primary cell walls of grasses, were in a lower energy configuration compared to wild type, suggesting a slightly weaker primary wall in MLG deficient mesophyll cells. Finally, taken together, these results suggest that MLG may influence cellulose deposition in mesophyll cell walls without significantly affecting anisotropic growth thus challenging MLG importance in cell wall expansion.« less

The Complex Cell Wall Composition of Syncytia Induced by Plant Parasitic Cyst Nematodes Reflects Both Function and Host Plant

PubMed Central

Zhang, Li; Lilley, Catherine J.; Imren, Mustafa; Knox, J. Paul; Urwin, Peter E.

2017-01-01

Plant–parasitic cyst nematodes induce the formation of specialized feeding structures, syncytia, within their host roots. These unique plant organs serve as the sole nutrient resource for development and reproduction throughout the biotrophic interaction. The multinucleate syncytium, which arises through local dissolution of cell walls and protoplast fusion of multiple adjacent cells, has dense cytoplasm containing numerous organelles, surrounded by thickened outer cell walls that must withstand high turgor pressure. However, little is known about how the constituents of the syncytial cell wall and their conformation support its role during nematode parasitism. We used a set of monoclonal antibodies, targeted to a range of plant cell wall components, to reveal the microstructures of syncytial cell walls induced by four of the most economically important cyst nematode species, Globodera pallida, Heterodera glycines, Heterodera avenae and Heterodera filipjevi, in their respective potato, soybean, and spring wheat host roots. In situ fluorescence analysis revealed highly similar cell wall composition of syncytia induced by G. pallida and H. glycines. Both consisted of abundant xyloglucan, methyl-esterified homogalacturonan and pectic arabinan. In contrast, the walls of syncytia induced in wheat roots by H. avenae and H. filipjevi contain little xyloglucan but are rich in feruloylated xylan and arabinan residues, with variable levels of mixed-linkage glucan. The overall chemical composition of syncytial cell walls reflected the general features of root cell walls of the different host plants. We relate specific components of syncytial cell walls, such as abundant arabinan, methyl-esterification status of pectic homogalacturonan and feruloylation of xylan, to their potential roles in forming a network to support both the strength and flexibility required for syncytium function. PMID:28680436

Multiple cell radiation detector system, and method, and submersible sonde

DOEpatents

Johnson, Larry O.; McIsaac, Charles V.; Lawrence, Robert S.; Grafwallner, Ervin G.

2002-01-01

A multiple cell radiation detector includes a central cell having a first cylindrical wall providing a stopping power less than an upper threshold; an anode wire suspended along a cylindrical axis of the central cell; a second cell having a second cylindrical wall providing a stopping power greater than a lower threshold, the second cylindrical wall being mounted coaxially outside of the first cylindrical wall; a first end cap forming a gas-tight seal at first ends of the first and second cylindrical walls; a second end cap forming a gas-tight seal at second ends of the first and second cylindrical walls; and a first group of anode wires suspended between the first and second cylindrical walls.

Transmission electron microscopy, fluorescence microscopy, and confocal raman microscopic analysis of ultrastructural and compositional heterogeneity of Cornus alba L. wood cell wall.

PubMed

Ma, Jianfeng; Ji, Zhe; Zhou, Xia; Zhang, Zhiheng; Xu, Feng

2013-02-01

Transmission electron microscopy (TEM), fluorescence microscopy, and confocal Raman microscopy can be used to characterize ultrastructural and compositional heterogeneity of plant cell walls. In this study, TEM observations revealed the ultrastructural characterization of Cornus alba L. fiber, vessel, axial parenchyma, ray parenchyma, and pit membrane between cells, notably with the ray parenchyma consisting of two well-defined layers. Fluorescence microscopy evidenced that cell corner middle lamella was more lignified than adjacent compound middle lamella and secondary wall with variation in lignification level from cell to cell. In situ Raman images showed that the inhomogeneity in cell wall components (cellulose and lignin) among different cells and within morphologically distinct cell wall layers. As the significant precursors of lignin biosynthesis, the pattern of coniferyl alcohol and aldehyde (joint abbreviation Lignin-CAA for both structures) distribution in fiber cell wall was also identified by Raman images, with higher concentration occurring in the fiber secondary wall where there was the highest cellulose concentration. Moreover, noteworthy was the observation that higher concentration of lignin and very minor amounts of cellulose were visualized in the pit membrane areas. These complementary microanalytical methods provide more accurate and complete information with regard to ultrastructural and compositional characterization of plant cell walls.

Overexpression of PhEXPA1 increases cell size, modifies cell wall polymer composition and affects the timing of axillary meristem development in Petunia hybrida.

PubMed

Zenoni, Sara; Fasoli, Marianna; Tornielli, Giovanni Battista; Dal Santo, Silvia; Sanson, Andrea; de Groot, Peter; Sordo, Sara; Citterio, Sandra; Monti, Francesca; Pezzotti, Mario

2011-08-01

• Expansins are cell wall proteins required for cell enlargement and cell wall loosening during many developmental processes. The involvement of the Petunia hybrida expansin A1 (PhEXPA1) gene in cell expansion, the control of organ size and cell wall polysaccharide composition was investigated by overexpressing PhEXPA1 in petunia plants. • PhEXPA1 promoter activity was evaluated using a promoter-GUS assay and the protein's subcellular localization was established by expressing a PhEXPA1-GFP fusion protein. PhEXPA1 was overexpressed in transgenic plants using the cauliflower mosaic virus (CaMV) 35S promoter. Fourier transform infrared (FTIR) and chemical analysis were used for the quantitative analysis of cell wall polymers. • The GUS and GFP assays demonstrated that PhEXPA1 is present in the cell walls of expanding tissues. The constitutive overexpression of PhEXPA1 significantly affected expansin activity and organ size, leading to changes in the architecture of petunia plants by initiating premature axillary meristem outgrowth. Moreover, a significant change in cell wall polymer composition in the petal limbs of transgenic plants was observed. • These results support a role for expansins in the determination of organ shape, in lateral branching, and in the variation of cell wall polymer composition, probably reflecting a complex role in cell wall metabolism. © 2011 The Authors. New Phytologist © 2011 New Phytologist Trust.

Tissue and cell-type co-expression networks of transcription factors and wood component genes in Populus trichocarpa.

PubMed

Shi, Rui; Wang, Jack P; Lin, Ying-Chung; Li, Quanzi; Sun, Ying-Hsuan; Chen, Hao; Sederoff, Ronald R; Chiang, Vincent L

2017-05-01

Co-expression networks based on transcriptomes of Populus trichocarpa major tissues and specific cell types suggest redundant control of cell wall component biosynthetic genes by transcription factors in wood formation. We analyzed the transcriptomes of five tissues (xylem, phloem, shoot, leaf, and root) and two wood forming cell types (fiber and vessel) of Populus trichocarpa to assemble gene co-expression subnetworks associated with wood formation. We identified 165 transcription factors (TFs) that showed xylem-, fiber-, and vessel-specific expression. Of these 165 TFs, 101 co-expressed (correlation coefficient, r > 0.7) with the 45 secondary cell wall cellulose, hemicellulose, and lignin biosynthetic genes. Each cell wall component gene co-expressed on average with 34 TFs, suggesting redundant control of the cell wall component gene expression. Co-expression analysis showed that the 101 TFs and the 45 cell wall component genes each has two distinct groups (groups 1 and 2), based on their co-expression patterns. The group 1 TFs (44 members) are predominantly xylem and fiber specific, and are all highly positively co-expressed with the group 1 cell wall component genes (30 members), suggesting their roles as major wood formation regulators. Group 1 TFs include a lateral organ boundary domain gene (LBD) that has the highest number of positively correlated cell wall component genes (36) and TFs (47). The group 2 TFs have 57 members, including 14 vessel-specific TFs, and are generally less correlated with the cell wall component genes. An exception is a vessel-specific basic helix-loop-helix (bHLH) gene that negatively correlates with 20 cell wall component genes, and may function as a key transcriptional suppressor. The co-expression networks revealed here suggest a well-structured transcriptional homeostasis for cell wall component biosynthesis during wood formation.

THESEUS 1, FERONIA and relatives: a family of cell wall-sensing receptor kinases?

PubMed

Cheung, Alice Y; Wu, Hen-Ming

2011-12-01

The plant cell wall provides form and integrity to the cell as well as a dynamic interface between a cell and its environment. Therefore mechanisms capable of policing changes in the cell wall, signaling cellular responses including those that would feedback regulate cell wall properties are expected to play important roles in facilitating growth and ensuring survival. Discoveries in the last few years that the Arabidopsis THESEUS 1 receptor-like kinase (RLK) may function as a sensor for cell wall defects to regulate growth and that its relatives FERONIA and ANXURs regulate pollen tube integrity imply strongly that they play key roles in cell wall-related processes. Furthermore, FERONIA acts as a cell surface regulator for RAC/ROP GTPases and activates production of reactive oxygen species which are, respectively, important molecular switches and mediators for diverse processes. These findings position the THESEUS 1/FERONIA family RLKs as surface regulators and potential cell wall sensors capable of broadly and profoundly impacting cellular pathways in response to diverse signals. Copyright © 2011 Elsevier Ltd. All rights reserved.

Characterization of xylan in the early stages of secondary cell wall formation in tobacco bright yellow-2 cells.

PubMed

Ishii, Tadashi; Matsuoka, Keita; Ono, Hiroshi; Ohnishi-Kameyama, Mayumi; Yaoi, Katsuro; Nakano, Yoshimi; Ohtani, Misato; Demura, Taku; Iwai, Hiroaki; Satoh, Shinobu

2017-11-15

The major polysaccharides present in the primary and secondary walls surrounding plant cells have been well characterized. However, our knowledge of the early stages of secondary wall formation is limited. To address this, cell walls were isolated from differentiating xylem vessel elements of tobacco bright yellow-2 (BY-2) cells induced by VASCULAR-RELATED NAC-DOMAIN7 (VND7). The walls of induced VND7-VP16-GR BY-2 cells consisted of cellulose, pectic polysaccharides, hemicelluloses, and lignin, and contained more xylan and cellulose compared with non-transformed BY-2 and uninduced VND7-VP16-GR BY-2 cells. A reducing end sequence of xylan containing rhamnose and galaturonic acid- residues is present in the walls of induced, uninduced, and non-transformed BY-2 cells. Glucuronic acid residues in xylan from walls of induced cells are O-methylated, while those of xylan in non-transformed BY-2 and uninduced cells are not. Our results show that xylan changes in chemical structure and amounts during the early stages of xylem differentiation. Copyright © 2017 Elsevier Ltd. All rights reserved.

The Modification of Cell Wall Properties by Expression of Recombinant Resilin in Transgenic Plants.

PubMed

Preis, Itan; Abramson, Miron; Shoseyov, Oded

2018-04-01

Plant tissue is composed of many different types of cells. Plant cells required to withstand mechanical pressure, such as vessel elements and fibers, have a secondary cell wall consisting of polysaccharides and lignin, which strengthen the cell wall structure and stabilize the cell shape. Previous attempts to alter the properties of the cell wall have mainly focused on reducing the amount of lignin or altering its structure in order to ease its extraction from raw woody materials for the pulp and paper and biorefinery industries. In this work, we propose the in vivo modification of the cell wall structure and mechanical properties by the introduction of resilin, an elastic protein that is able to crosslink with lignin monomers during cell wall synthesis. The effects of resilin were studied in transgenic eucalyptus plants. The protein was detected within the cell wall and its expression led to an increase in the elastic modulus of transgenic stems. In addition, transgenic stems displayed a higher yield point and toughness, indicating that they were able to absorb more energy before breaking.

Deformation and failure mechanism of secondary cell wall in Spruce late wood

NASA Astrophysics Data System (ADS)

Adusumalli, Ramesh-Babu; Raghavan, Rejin; Ghisleni, Rudy; Zimmermann, Tanja; Michler, Johann

2010-08-01

The deformation and failure of the secondary cell wall of Spruce wood was studied by in-situ SEM compression of micropillars machined by the focused ion beam technique. The cell wall exhibited yield strength values of approximately 160 MPa and large scale plasticity. High resolution SEM imaging post compression revealed bulging of the pillars followed by shear failure. With additional aid of cross-sectional analysis of the micropillars post compression, a model for deformation and failure mechanism of the cell wall has been proposed. The cell wall consists of oriented cellulose microfibrils with high aspect ratio embedded in a hemicellulose-lignin matrix. The deformation of the secondary wall occurs by asymmetric out of plane bulging because of buckling of the microfibrils. Failure of the cell wall following the deformation occurs by the formation of a shear or kink band.

Cell Wall Localization of Two DUF642 Proteins, BIIDXI and TEEBE, during Meloidogyne incognita Early Inoculation

PubMed Central

Salazar-Iribe, Alexis; Zúñiga-Sánchez, Esther; Mejía, Emma Zavaleta; Gamboa-deBuen, Alicia

2017-01-01

The root-knot nematode Meloidogyne incognita infects a variety of plants, including Arabidopsis thaliana. During migration, root-knot nematodes secrete different proteins to modify cell walls, which include pectolytic enzymes. However, the contribution of host cell wall proteins has not been described during this process. The function of two DUF642 cell wall proteins, BIIDXI (BDX, At4g32460) and TEEBE (TEB, At2g41800), in plant development could be related to the regulation of pectin methyl esterification status in the cell walls of different tissues. Accordingly, the expression of these two genes is up-regulated by auxin. BDX and TEB were highly induced during early M. incognita inoculation. Moreover, cell wall localization of the proteins was also induced. The cell wall localization of BDX and TEB DUF642 proteins during M. incognita early inoculation suggested that these two proteins could be involved in the regulation of the degree of pectin methylation during cell separation. PMID:29238286

Mechanism of cassava tuber cell wall weakening by dilute sodium hydroxide steeping.

PubMed

Odoch, Martin; Buys, Elna M; Taylor, John R N

2017-08-01

Steeping of cassava root pieces in 0.75% NaOH in combination with wet milling was investigated to determine whether and how dilute NaOH modifies cassava cell walls. Gas chromatography data of cell wall constituent sugar composition and Fourier transform infrared (FTIR) data showed that NaOH steeping reduced the level of pectin in cassava cell walls. FTIR and wide-angle X-ray scattering spectroscopy also indicated that NaOH steeping combined with fine milling slightly reduced cellulose crystallinity. Scanning electron microscopy showed that NaOH steeping produced micropores in the cell walls and light microscopy revealed that NaOH steeping increased disaggregation of parenchyma cells. Steeping of ground cassava in NaOH resulted in a 12% decrease in large residue particles and approx. 4% greater starch yield with wet milling. Therefore dilute NaOH steeping can improve the effectiveness of wet milling in disintegrating cell walls through solubilisation of pectin, thereby reduced cell wall strength. Copyright © 2017 Elsevier Ltd. All rights reserved.

Phenolic components of the primary cell wall. Feruloylated disaccharides of D-galactose and L-arabinose from spinach polysaccharide.

PubMed Central

Fry, S C

1982-01-01

1. Cell walls from rapidly growing cell suspension cultures of Spinacia oleracea L. contained ferulic acid and p-coumaric acid esterified with a water-insoluble polymer. 2. Prolonged treatment with trypsin did not release may feruloyl esters from dearabinofuranosylated cell walls, and the polymer was also insoluble in phenol/acetic acid/water (2:1:1, w/v/v). 3. Treatment of the cell walls with the fungal hydrolase preparation "Driselase' did liberate low-Mr feruloyl esters. The major esters were 4-O-(6-O-feruloyl-beta-D-galactopyranosyl)-D-galactose and 3?-O-feruloyl-alpha-L-arabinopyranosyl)-L-arabinose. These two esters accounted for about 60% of the cell-wall ferulate. 4. It is concluded that the feruloylation of cell-wall polymers is not a random process, but occurs at very specific sites, probably on the arabinogalactan component of pectin. 5. The possible role of such phenolic substituents in cell-wall architecture and growth is discussed. PMID:7115300

Shifting foundations: the mechanical cell wall and development.

PubMed

Braybrook, Siobhan A; Jönsson, Henrik

2016-02-01

The cell wall has long been acknowledged as an important physical mediator of growth in plants. Recent experimental and modelling work has brought the importance of cell wall mechanics into the forefront again. These data have challenged existing dogmas that relate cell wall structure to cell/organ growth, that uncouple elasticity from extensibility, and those which treat the cell wall as a passive and non-stressed material. Within this review we describe experiments and models which have changed the ways in which we view the mechanical cell wall, leading to new hypotheses and research avenues. It has become increasingly apparent that while we often wish to simplify our systems, we now require more complex multi-scale experiments and models in order to gain further insight into growth mechanics. We are currently experiencing an exciting and challenging shift in the foundations of our understanding of cell wall mechanics in growth and development. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

30 years of battling the cell wall.

PubMed

Latgé, J P

2017-01-01

In Aspergillus fumigatus, like in other pathogenic fungi, the cell wall is essential for fungal growth as well as for resisting environmental stresses such as phagocytic killing. Most of the chemical analyses undertaken on the cell wall of A. fumigatus are focused on the mycelial cell wall because it is the vegetative stage of the fungus. However, the cell walls of the mycelium and conidium (which is the infective propagule) are different especially at the level of the surface layer, which plays a significant role in the interaction between A. fumigatus conidia and phagocytic cells of the immune system. In spite of the essential function of the cell wall in fungal life, progresses have been extremely slow in the understanding of biosynthesis as well in the identification of the key host responses against the cell wall components. A major difficulty is the fact that the composition and structural organization of the cell wall is not immutably set and is constantly reshuffled depending on the environmental conditions. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Discovery of novel cell wall-active compounds using P ywaC, a sensitive reporter of cell wall stress, in the model gram-positive bacterium Bacillus subtilis.

PubMed

Czarny, T L; Perri, A L; French, S; Brown, E D

2014-06-01

The emergence of antibiotic resistance in recent years has radically reduced the clinical efficacy of many antibacterial treatments and now poses a significant threat to public health. One of the earliest studied well-validated targets for antimicrobial discovery is the bacterial cell wall. The essential nature of this pathway, its conservation among bacterial pathogens, and its absence in human biology have made cell wall synthesis an attractive pathway for new antibiotic drug discovery. Herein, we describe a highly sensitive screening methodology for identifying chemical agents that perturb cell wall synthesis, using the model of the Gram-positive bacterium Bacillus subtilis. We report on a cell-based pilot screen of 26,000 small molecules to look for cell wall-active chemicals in real time using an autonomous luminescence gene cluster driven by the promoter of ywaC, which encodes a guanosine tetra(penta)phosphate synthetase that is expressed under cell wall stress. The promoter-reporter system was generally much more sensitive than growth inhibition testing and responded almost exclusively to cell wall-active antibiotics. Follow-up testing of the compounds from the pilot screen with secondary assays to verify the mechanism of action led to the discovery of 9 novel cell wall-active compounds. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

14 CFR 121.245 - Fire walls.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 14 Aeronautics and Space 3 2010-01-01 2010-01-01 false Fire walls. 121.245 Section 121.245 Aeronautics and Space FEDERAL AVIATION ADMINISTRATION, DEPARTMENT OF TRANSPORTATION (CONTINUED) AIR CARRIERS..., FLAG, AND SUPPLEMENTAL OPERATIONS Special Airworthiness Requirements § 121.245 Fire walls. Each engine...

Investigation of the functional role of CSLD proteins in plant cell wall deposition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nielsen, Erik Etlar

The overall goal of this research proposal was to characterize the molecular machinery responsible for polarized secretion of cell wall components in Arabidopsis thaliana. We have used the polarized expansion that occurs during root hair cell growth to identify membrane trafficking pathways involved in polarized secretion of cell wall components to the expanding tips of these cells, and we have recently shown that CSLD3 is preferentially targeted to the apical plasma membranes in root hair cells, where it plays essential roles during cell wall deposition in these cells. The specific aims of the project are designed to answer the followingmore » objective: Identification of the cell wall polysaccharide class that CSLD proteins synthesize.« less

Characterization and elimination of undesirable protein residues in plant cell walls for enhancing lignin analysis by solution-state 2D gel-NMR methods

USDA-ARS?s Scientific Manuscript database

Proteins exist in every plant cell wall. Certain protein residues interfere with lignin characterization and quantification. The current solution-state 2D-NMR technique (gel-NMR) for whole plant cell wall structural profiling provides detailed information regarding cell walls and proteins. However, ...

Cell Wall Structure in Cells Adapted to Growth on the Cellulose-Synthesis Inhibitor 2,6-Dichlorobenzonitrile 1

PubMed Central

Shedletzky, Esther; Shmuel, Miri; Trainin, Tali; Kalman, Sara; Delmer, Deborah

1992-01-01

Our previous work (E. Shedletzky, M. Shmuel, D.P. Delmer, D.T.A. Lamport [1990] Plant Physiol 94:980-987) showed that suspension-cultured tomato cells adapted to growth on the cellulose synthesis inhibitor 2,6-dichlorobenzonitrile (DCB) have a markedly altered cell wall composition, most notably a markedly reduced level of the cellulose-xyloglucan network. This study compares the adaptation to DCB of two cell lines from dicots (tomato [Lycopersicon esculentum] and tobacco [Nicotiana tabacum]) and a Graminaceous monocot (barley [Hordeum bulbosum] endosperm). The difference in wall structures between the dicots and the monocot is reflected in the very different types of wall modifications induced by growth on DCB. The dicots, having reduced levels of cellulose and xyloglucan, possess walls the major integrity of which is provided by Ca2+-bridged pectates because protoplasts can be prepared from these cells simply by treatment with divalent cation chelator and a purified endopolygalacturonase. The tensile strength of these walls is considerably less than walls from nonadapted cells, but wall porosity is not altered. In contrast, walls from adapted barley cells contain very little pectic material and normal to elevated levels of noncellulosic polysaccharides compared with walls from nonadapted cells. Surprisingly, they have tensile strengths higher than their nonadapted counterpart, although cellulose levels are reduced by 70%. Evidence is presented that these walls obtain their additional strength by an altered pattern of cross-linking of polymers involving phenolic components. Such cross-linking may also explain the observation that the porosity of these walls is also considerably reduced. Cells of adapted lines of both the dicots and barley are resistant to plasmolysis, suggesting that they possess very strong connections between the wall and the plasma membrane. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:16652933

Engineering increased thermostability in the thermostable GH-11 xylanase from Thermobacillus xylanilyticus.

PubMed

Paës, Gabriel; O'Donohue, Michael J

2006-09-18

Enzymatic hydrolysis constitutes an attractive strategy for biorefining of abundant, low-cost agricultural by-products such as wheat bran and straw. However, to adopt such an approach, efficient enzymes are required, in particular xylanases. To promote heat-induced disorganization of the complex cell wall network in wheat bran and thus increase enzymatic hydrolysis, we have attempted to improve the thermoresistance of a GH-11 xylanase that is already moderately thermostable. Using a previously described engineering strategy that involves the introduction of disulphide bridges, a mutant (Tx-xyl-SS3) displaying enhanced thermostability and thermoactivity was obtained. The half life at 70 degrees C (180 min) of Tx-xyl-SS3 is 10-fold greater than that of the wild type enzyme and its specific activity is almost doubled (3500 IU mg(-1)). Despite these improvements, Tx-xyl-SS3 was unsuitable for use at significantly higher reaction temperatures (i.e. 85-95 degrees C) and thus the initial objective of this study remained unaccomplished. However, unexpectedly even at the normal hydrolytic temperature (60 degrees C), Tx-xyl-SS3 was able to solubilize 50% of the wheat bran arabinoxylans, 10 points more than the wild type enzyme in parallel reactions. The data presented here show that this improvement is not directly linked to the increase in thermostability and/or thermoactivity, but rather to other unidentified changes to physico-chemical properties that may allow Tx-xyl-SS3 to better penetrate the cell wall network in wheat bran.

Attachment of Salmonella strains to a plant cell wall model is modulated by surface characteristics and not by specific carbohydrate interactions.

PubMed

Tan, Michelle Sze-Fan; Moore, Sean C; Tabor, Rico F; Fegan, Narelle; Rahman, Sadequr; Dykes, Gary A

2016-09-15

Processing of fresh produce exposes cut surfaces of plant cell walls that then become vulnerable to human foodborne pathogen attachment and contamination, particularly by Salmonella enterica. Plant cell walls are mainly composed of the polysaccharides cellulose, pectin and hemicelluloses (predominantly xyloglucan). Our previous work used bacterial cellulose-based plant cell wall models to study the interaction between Salmonella and the various plant cell wall components. We demonstrated that Salmonella attachment was favoured in the presence of pectin while xyloglucan had no effect on its attachment. Xyloglucan significantly increased the attachment of Salmonella cells to the plant cell wall model only when it was in association with pectin. In this study, we investigate whether the plant cell wall polysaccharides mediate Salmonella attachment to the bacterial cellulose-based plant cell wall models through specific carbohydrate interactions or through the effects of carbohydrates on the physical characteristics of the attachment surface. We found that none of the monosaccharides that make up the plant cell wall polysaccharides specifically inhibit Salmonella attachment to the bacterial cellulose-based plant cell wall models. Confocal laser scanning microscopy showed that Salmonella cells can penetrate and attach within the tightly arranged bacterial cellulose network. Analysis of images obtained from atomic force microscopy revealed that the bacterial cellulose-pectin-xyloglucan composite with 0.3 % (w/v) xyloglucan, previously shown to have the highest number of Salmonella cells attached to it, had significantly thicker cellulose fibrils compared to other composites. Scanning electron microscopy images also showed that the bacterial cellulose and bacterial cellulose-xyloglucan composites were more porous when compared to the other composites containing pectin. Our study found that the attachment of Salmonella cells to cut plant cell walls was not mediated by specific carbohydrate interactions. This suggests that the attachment of Salmonella strains to the plant cell wall models were more dependent on the structural characteristics of the attachment surface. Pectin reduces the porosity and space between cellulose fibrils, which then forms a matrix that is able to retain Salmonella cells within the bacterial cellulose network. When present with pectin, xyloglucan provides a greater surface for Salmonella cells to attach through the thickening of cellulose fibrils.

Changes in Cell Wall Polysaccharides Associated With Growth 1

PubMed Central

Nevins, Donald J.; English, Patricia D.; Albersheim, Peter

1968-01-01

Changes in the polysaccharide composition of Phaseolus vulgaris, P. aureus, and Zea mays cell walls were studied during the first 28 days of seedling development using a gas chromatographic method for the analysis of neutral sugars. Acid hydrolysis of cell wall material from young tissues liberates rhamnose, fucose, arabinose, xylose, mannose, galactose, and glucose which collectively can account for as much as 70% of the dry weight of the wall. Mature walls in fully expanded tissues of these same plants contain less of these constituents (10%-20% of dry wt). Gross differences are observed between developmental patterns of the cell wall in the various parts of a seedling, such as root, stem, and leaf. The general patterns of wall polysaccharide composition change, however, are similar for analogous organs among the varieties of a species. Small but significant differences in the rates of change in sugar composition were detected between varieties of the same species which exhibited different growth patterns. The cell walls of species which are further removed phylogenetically exhibit even more dissimilar developmental patterns. The results demonstrate the dynamic nature of the cell wall during growth as well as the quantitative and qualitative exactness with which the biosynthesis of plant cell walls is regulated. PMID:16656862

A plant cell division algorithm based on cell biomechanics and ellipse-fitting.

PubMed

Abera, Metadel K; Verboven, Pieter; Defraeye, Thijs; Fanta, Solomon Workneh; Hertog, Maarten L A T M; Carmeliet, Jan; Nicolai, Bart M

2014-09-01

The importance of cell division models in cellular pattern studies has been acknowledged since the 19th century. Most of the available models developed to date are limited to symmetric cell division with isotropic growth. Often, the actual growth of the cell wall is either not considered or is updated intermittently on a separate time scale to the mechanics. This study presents a generic algorithm that accounts for both symmetrically and asymmetrically dividing cells with isotropic and anisotropic growth. Actual growth of the cell wall is simulated simultaneously with the mechanics. The cell is considered as a closed, thin-walled structure, maintained in tension by turgor pressure. The cell walls are represented as linear elastic elements that obey Hooke's law. Cell expansion is induced by turgor pressure acting on the yielding cell-wall material. A system of differential equations for the positions and velocities of the cell vertices as well as for the actual growth of the cell wall is established. Readiness to divide is determined based on cell size. An ellipse-fitting algorithm is used to determine the position and orientation of the dividing wall. The cell vertices, walls and cell connectivity are then updated and cell expansion resumes. Comparisons are made with experimental data from the literature. The generic plant cell division algorithm has been implemented successfully. It can handle both symmetrically and asymmetrically dividing cells coupled with isotropic and anisotropic growth modes. Development of the algorithm highlighted the importance of ellipse-fitting to produce randomness (biological variability) even in symmetrically dividing cells. Unlike previous models, a differential equation is formulated for the resting length of the cell wall to simulate actual biological growth and is solved simultaneously with the position and velocity of the vertices. The algorithm presented can produce different tissues varying in topological and geometrical properties. This flexibility to produce different tissue types gives the model great potential for use in investigations of plant cell division and growth in silico.

A glycosylphosphatidylinositol anchor is required for membrane localization but dispensable for cell wall association of chitin deacetylase 2 in Cryptococcus neoformans.

PubMed

Gilbert, Nicole M; Baker, Lorina G; Specht, Charles A; Lodge, Jennifer K

2012-01-01

Cell wall proteins (CWPs) mediate important cellular processes in fungi, including adhesion, invasion, biofilm formation, and flocculation. The current model of fungal cell wall organization includes a major class of CWPs covalently bound to β-1,6-glucan via a remnant of a glycosylphosphatidylinositol (GPI) anchor. This model was established by studies of ascomycetes more than a decade ago, and relatively little work has been done with other fungi, although the presumption has been that proteins identified in the cell wall which contain a predicted GPI anchor are covalently linked to cell wall glucans. The pathogenic basidiomycete Cryptococcus neoformans encodes >50 putatively GPI-anchored proteins, some of which have been identified in the cell wall. One of these proteins is chitin deacetylase 2 (Cda2), an enzyme responsible for converting chitin to chitosan, a cell wall polymer recently established as a virulence factor for C. neoformans infection of mammalian hosts. Using a combination of biochemistry, molecular biology, and genetics, we show that Cda2 is GPI anchored to membranes but noncovalently associated with the cell wall by means independent of both its GPI anchor and β-1,6-glucan. We also show that Cda2 produces chitosan when localized to the plasma membrane, but association with the cell wall is not essential for this process, thereby providing insight into the mechanism of chitosan biosynthesis. These results increase our understanding of the surface of C. neoformans and provide models of cell walls likely applicable to other undercharacterized basidiomycete pathogenic fungi. The surface of a pathogenic microbe is a major interface with its host. In fungi, the outer surface consists of a complex matrix known as the cell wall, which includes polysaccharides, proteins, and other molecules. The mammalian host recognizes many of these surface molecules and mounts appropriate responses to combat the microbial infection. Cryptococcus neoformans is a serious fungal pathogen that kills over 600,000 people annually. It converts most of its chitin, a cell wall polysaccharide, to chitosan, which is necessary for virulence. Chitin deacetylase enzymes have been identified in the cell wall, and our studies were undertaken to understand how the deacetylase is linked to the wall and where it has activity. Our results have implications for the current model of chitosan biosynthesis and further challenge the paradigm of covalent linkages between cell wall proteins and polysaccharides through a lipid modification of the protein.

Reciprocal Interactions between Cadmium-Induced Cell Wall Responses and Oxidative Stress in Plants

PubMed Central

Loix, Christophe; Huybrechts, Michiel; Vangronsveld, Jaco; Gielen, Marijke; Keunen, Els; Cuypers, Ann

2017-01-01

Cadmium (Cd) pollution renders many soils across the world unsuited or unsafe for food- or feed-orientated agriculture. The main mechanism of Cd phytotoxicity is the induction of oxidative stress, amongst others through the depletion of glutathione. Oxidative stress can damage lipids, proteins, and nucleic acids, leading to growth inhibition or even cell death. The plant cell has a variety of tools to defend itself against Cd stress. First and foremost, cell walls might prevent Cd from entering and damaging the protoplast. Both the primary and secondary cell wall have an array of defensive mechanisms that can be adapted to cope with Cd. Pectin, which contains most of the negative charges within the primary cell wall, can sequester Cd very effectively. In the secondary cell wall, lignification can serve to immobilize Cd and create a tougher barrier for entry. Changes in cell wall composition are, however, dependent on nutrients and conversely might affect their uptake. Additionally, the role of ascorbate (AsA) as most important apoplastic antioxidant is of considerable interest, due to the fact that oxidative stress is a major mechanism underlying Cd toxicity, and that AsA biosynthesis shares several links with cell wall construction. In this review, modifications of the plant cell wall in response to Cd exposure are discussed. Focus lies on pectin in the primary cell wall, lignification in the secondary cell wall and the importance of AsA in the apoplast. Regarding lignification, we attempt to answer the question whether increased lignification is merely a consequence of Cd toxicity, or rather an elicited defense response. We propose a model for lignification as defense response, with a central role for hydrogen peroxide as substrate and signaling molecule. PMID:29163592

Genetic modification of plant cell walls to enhance biomass yield and biofuel production in bioenergy crops.

PubMed

Wang, Yanting; Fan, Chunfen; Hu, Huizhen; Li, Ying; Sun, Dan; Wang, Youmei; Peng, Liangcai

2016-01-01

Plant cell walls represent an enormous biomass resource for the generation of biofuels and chemicals. As lignocellulose property principally determines biomass recalcitrance, the genetic modification of plant cell walls has been posed as a powerful solution. Here, we review recent progress in understanding the effects of distinct cell wall polymers (cellulose, hemicelluloses, lignin, pectin, wall proteins) on the enzymatic digestibility of biomass under various physical and chemical pretreatments in herbaceous grasses, major agronomic crops and fast-growing trees. We also compare the main factors of wall polymer features, including cellulose crystallinity (CrI), hemicellulosic Xyl/Ara ratio, monolignol proportion and uronic acid level. Furthermore, the review presents the main gene candidates, such as CesA, GH9, GH10, GT61, GT43 etc., for potential genetic cell wall modification towards enhancing both biomass yield and enzymatic saccharification in genetic mutants and transgenic plants. Regarding cell wall modification, it proposes a novel groove-like cell wall model that highlights to increase amorphous regions (density and depth) of the native cellulose microfibrils, providing a general strategy for bioenergy crop breeding and biofuel processing technology. Copyright © 2016 Elsevier Inc. All rights reserved.

Arabidopsis thaliana plants lacking the ARP2/3 complex show defects in cell wall assembly and auxin distribution.

PubMed

Pratap Sahi, Vaidurya; Cifrová, Petra; García-González, Judith; Kotannal Baby, Innu; Mouillé, Gregory; Gineau, Emilie; Müller, Karel; Baluška, František; Soukup, Aleš; Petrášek, Jan; Schwarzerová, Katerina

2017-12-25

The cytoskeleton plays an important role in the synthesis of plant cell walls. Both microtubules and actin cytoskeleton are known to be involved in the morphogenesis of plant cells through their role in cell wall building. The role of ARP2/3-nucleated actin cytoskeleton in the morphogenesis of cotyledon pavement cells has been described before. Seedlings of Arabidopsis mutants lacking a functional ARP2/3 complex display specific cell wall-associated defects. In three independent Arabidopsis mutant lines lacking subunits of the ARP2/3 complex, phenotypes associated with the loss of the complex were analysed throughout plant development. Organ size and anatomy, cell wall composition, and auxin distribution were investigated. ARP2/3-related phenotype is associated with changes in cell wall composition, and the phenotype is manifested especially in mature tissues. Cell walls of mature plants contain less cellulose and a higher amount of homogalacturonan, and display changes in cell wall lignification. Vascular bundles of mutant inflorescence stems show a changed pattern of AUX1-YFP expression. Plants lacking a functional ARP2/3 complex have decreased basipetal auxin transport. The results suggest that the ARP2/3 complex has a morphogenetic function related to cell wall synthesis and auxin transport. © The Author(s) 2017. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Local differentiation of cell wall matrix polysaccharides in sinuous pavement cells: its possible involvement in the flexibility of cell shape.

PubMed

Sotiriou, P; Giannoutsou, E; Panteris, E; Galatis, B; Apostolakos, P

2018-03-01

The distribution of homogalacturonans (HGAs) displaying different degrees of esterification as well as of callose was examined in cell walls of mature pavement cells in two angiosperm and two fern species. We investigated whether local cell wall matrix differentiation may enable pavement cells to respond to mechanical tension forces by transiently altering their shape. HGA epitopes, identified with 2F4, JIM5 and JIM7 antibodies, and callose were immunolocalised in hand-made or semithin leaf sections. Callose was also stained with aniline blue. The structure of pavement cells was studied with light and transmission electron microscopy (TEM). In all species examined, pavement cells displayed wavy anticlinal cell walls, but the waviness pattern differed between angiosperms and ferns. The angiosperm pavement cells were tightly interconnected throughout their whole depth, while in ferns they were interconnected only close to the external periclinal cell wall and intercellular spaces were developed between them close to the mesophyll. Although the HGA epitopes examined were located along the whole cell wall surface, the 2F4- and JIM5- epitopes were especially localised at cell lobe tips. In fern pavement cells, the contact sites were impregnated with callose and JIM5-HGA epitopes. When tension forces were applied on leaf regions, the pavement cells elongated along the stretching axis, due to a decrease in waviness of anticlinal cell walls. After removal of tension forces, the original cell shape was resumed. The presented data support that HGA epitopes make the anticlinal pavement cell walls flexible, in order to reversibly alter their shape. Furthermore, callose seems to offer stability to cell contacts between pavement cells, as already suggested in photosynthetic mesophyll cells. © 2017 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.

Exploring the Role of Cell Wall-Related Genes and Polysaccharides during Plant Development.

PubMed

Tucker, Matthew R; Lou, Haoyu; Aubert, Matthew K; Wilkinson, Laura G; Little, Alan; Houston, Kelly; Pinto, Sara C; Shirley, Neil J

2018-05-31

The majority of organs in plants are not established until after germination, when pluripotent stem cells in the growing apices give rise to daughter cells that proliferate and subsequently differentiate into new tissues and organ primordia. This remarkable capacity is not only restricted to the meristem, since maturing cells in many organs can also rapidly alter their identity depending on the cues they receive. One general feature of plant cell differentiation is a change in cell wall composition at the cell surface. Historically, this has been viewed as a downstream response to primary cues controlling differentiation, but a closer inspection of the wall suggests that it may play a much more active role. Specific polymers within the wall can act as substrates for modifications that impact receptor binding, signal mobility, and cell flexibility. Therefore, far from being a static barrier, the cell wall and its constituent polysaccharides can dictate signal transmission and perception, and directly contribute to a cell's capacity to differentiate. In this review, we re-visit the role of plant cell wall-related genes and polysaccharides during various stages of development, with a particular focus on how changes in cell wall machinery accompany the exit of cells from the stem cell niche.

A new picture of cell wall protein dynamics in elongating cells of Arabidopsis thaliana: Confirmed actors and newcomers

PubMed Central

Irshad, Muhammad; Canut, Hervé; Borderies, Gisèle; Pont-Lezica, Rafael; Jamet, Elisabeth

2008-01-01

Background Cell elongation in plants requires addition and re-arrangements of cell wall components. Even if some protein families have been shown to play roles in these events, a global picture of proteins present in cell walls of elongating cells is still missing. A proteomic study was performed on etiolated hypocotyls of Arabidopsis used as model of cells undergoing elongation followed by growth arrest within a short time. Results Two developmental stages (active growth and after growth arrest) were compared. A new strategy consisting of high performance cation exchange chromatography and mono-dimensional electrophoresis was established for separation of cell wall proteins. This work allowed identification of 137 predicted secreted proteins, among which 51 had not been identified previously. Apart from expected proteins known to be involved in cell wall extension such as xyloglucan endotransglucosylase-hydrolases, expansins, polygalacturonases, pectin methylesterases and peroxidases, new proteins were identified such as proteases, proteins related to lipid metabolism and proteins of unknown function. Conclusion This work highlights the CWP dynamics that takes place between the two developmental stages. The presence of proteins known to be related to cell wall extension after growth arrest showed that these proteins may play other roles in cell walls. Finally, putative regulatory mechanisms of protein biological activity are discussed from this global view of cell wall proteins. PMID:18796151

[Molecular mechanism of AtGA3OX1 and AtGA3OX2 genes affecting secondary wall thickening in stems in Arabidopsis].

PubMed

Wang, Zeng-Guang; Chai, Guo-Hua; Wang, Zhi-Yao; Tang, Xian-Feng; Sun, Chang-Jiang; Zhou, Gong-Ke; Ma, San-Mei

2013-05-01

Bioactive gibberellins (GAs) are a type of important plant growth regulators, which play the key roles in multiple processes, such as seed germination, leaf expansion, flowering, fruit bearing, and stem development. Its biosynthesis is regulated by a variety of enzymes including gibberellin 3-oxidase that is a key rate-limiting enzyme. In Arabidopsis, gibberellin 3-oxidase consists of four members, of which AtGA3OX1 and AtGA3OX2 are highly expressed in stems, suggesting the potential roles in the stem development played by the two genes. To date, there are few studies on AtGA3OX1 and AtGA3OX2 regulating secondary wall thickening in stems. In this study, we used the atga3ox1atga3ox2 double mutant as the materials to study the effects of AtGA3OX1 and AtGA3OX2 genes on secondary wall thickening in stems. The results indicated that simulations repression of AtGA3OX1 and AtGA3OX2 genes resulted in significantly reduction of secondary wall thickening of fiber cells, but not that of vessel cells. Three main components (cellulose, hemicelluloses, and lignin) were also dramatically suppressed in the double mutants. qRT-PCR analysis demonstrated that the expressions of secondary wall biosynthetic genes and the associated transcription factors were obviously affected in AtGA3OX1 and AtGA3OX2 double mutant. Therefore, we presume that Arabidopsis AtGA3OX1 and AtGA3OX2 genes might activate the expression of these transcription factors, thus regulate secondary wall thickening in stems. Together, our results provide a theoretical basis for enhancing the lodging resistance of food crops and improving the biomass of energy plants by genetically engineering Arabidopsis AtGA3OX homologs.

Grass cell walls: A story of cross-linking

USDA-ARS?s Scientific Manuscript database

Cell wall matrices are complex composites mainly of polysaccharides, phenolics (monomers and polymers), and protein. We are beginning to understand the synthesis of these major wall components individually, but still have a poor understanding of how the cell wall components are assembled into comple...

Plant cell wall extensibility: connecting plant cell growth with cell wall structure, mechanics, and the action of wall-modifying enzymes.

PubMed

Cosgrove, Daniel J

2016-01-01

The advent of user-friendly instruments for measuring force/deflection curves of plant surfaces at high spatial resolution has resulted in a recent outpouring of reports of the 'Young's modulus' of plant cell walls. The stimulus for these mechanical measurements comes from biomechanical models of morphogenesis of meristems and other tissues, as well as single cells, in which cell wall stress feeds back to regulate microtubule organization, auxin transport, cellulose deposition, and future growth directionality. In this article I review the differences between elastic modulus and wall extensibility in the context of cell growth. Some of the inherent complexities, assumptions, and potential pitfalls in the interpretation of indentation force/deflection curves are discussed. Reported values of elastic moduli from surface indentation measurements appear to be 10- to >1000-fold smaller than realistic tensile elastic moduli in the plane of plant cell walls. Potential reasons for this disparity are discussed, but further work is needed to make sense of the huge range in reported values. The significance of wall stress relaxation for growth is reviewed and connected to recent advances and remaining enigmas in our concepts of how cellulose, hemicellulose, and pectins are assembled to make an extensible cell wall. A comparison of the loosening action of α-expansin and Cel12A endoglucanase is used to illustrate two different ways in which cell walls may be made more extensible and the divergent effects on wall mechanics. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Critical cell wall hole size for lysis in Gram-positive bacteria

NASA Astrophysics Data System (ADS)

Mitchell, Gabriel; Wiesenfeld, Kurt; Nelson, Daniel; Weitz, Joshua

2013-03-01

Gram-positive bacteria transport molecules necessary for their survival through holes in their cell wall. The holes in cell walls need to be large enough to let critical nutrients pass through. However, the cell wall must also function to prevent the bacteria's membrane from protruding through a large hole into the environment and lysing the cell. As such, we hypothesize that there exists a range of cell wall hole sizes that allow for molecule transport but prevent membrane protrusion. Here we develop and analyze a biophysical theory of the response of a Gram-positive cell's membrane to the formation of a hole in the cell wall. We predict a critical hole size in the range 15-24nm beyond which lysis occurs. To test our theory, we measured hole sizes in Streptococcus pyogenes cells undergoing enzymatic lysis via transmission electron microscopy. The measured hole sizes are in strong agreement with our theoretical prediction. Together, the theory and experiments provide a means to quantify the mechanisms of death of Gram-positive cells via enzymatically mediated lysis and provides insight into the range of cell wall hole sizes compatible with bacterial homeostasis.

Rare Earth Element Recovery from Low-Grade Feedstocks Using Engineered E. coli

NASA Astrophysics Data System (ADS)

Brewer, A. W.; Park, D.; Jiao, Y.

2017-12-01

Rare earth elements (REEs) are critical materials for emerging science and technology industries, especially in the field of clean energy. However, their supply is potentially at risk due to political and economic concerns. The exploitation of new, low-grade REE sources in the United States, such as geothermal brines and mine tailings, may help to mitigate that supply risk. To purify and concentrate REEs from these sources, we have developed a biosorption approach using engineered E. coli cells that express a lanthanide binding tag on the cell surface. This tag has a high selectivity for REEs that enhances the native cell wall adsorption properties; the terbium adsorption capacity was increased approximately 2-fold, and the REE surface affinity was increased compared to all non-REE metals except copper. This biosorption method offers advantages over conventional REE extraction methods as it is inexpensive, environmentally friendly, and effective with low-grade feedstocks. In order to expand this method to an industrial scale, the cells must be contained in a durable material that permits the cell surfaces to function in a variety of bioreactor systems and to be reused through multiple adsorption and desorption cycles. Polyethylene glycol diacrylate (PEGDA) beads, with diameters from 200-400 um, can be impregnated with high concentrations of cells, and show promise in the selective adsorption of REEs from solution. In the future, the application of the adsorptive qualities of these engineered cells may be expanded to include other valuable metals, such as indium and gallium, to further develop the economic potential of this approach. Prepared by LLNL under Contract DE-AC52-07NA27344. LLNL-ABS-736022.

Elevated Cell Wall Serine in Pleiotropic Staphylococcal Mutants

PubMed Central

Korman, Ruth Z.

1966-01-01

Korman, Ruth Z. (Cornell University, Ithaca, N.Y.). Elevated cell wall serine in pleiotropic staphylococcal mutants. J. Bacteriol. 92:762–768. 1966.—Physically purified cell walls were prepared from two staphylococcal strains and from pleiotropic variants derived from them. The quantitative amino acid and amino sugar content of these walls is reported. The pleiotypes, which are identified culturally by their failure to elaborate coagulase, their resistance to bacteriophage, and their sensitivity to mannitol, have altered molar ratios of amino acids and amino sugars in their cell walls. In comparison with lysine content, the serine content of the mutant wall is elevated and the glycine content is reduced. The glucosamine content is reduced also. It is postulated that the pleiotropic mutants possess an altered cell wall biosynthetic pathway. Images PMID:5922547

Cell wall integrity signaling in plants: "To grow or not to grow that's the question".

PubMed

Voxeur, Aline; Höfte, Herman

2016-09-01

Plants, like yeast, have the ability to monitor alterations in the cell wall architecture that occur during normal growth or in changing environments and to trigger compensatory changes in the cell wall. We discuss how recent advances in our understanding of the cell wall architecture provide new insights into the role of cell wall integrity sensing in growth control. Next we review the properties of membrane receptor-like kinases that have roles in pH control, mechano-sensing and reactive oxygen species accumulation in growing cells and which may be the plant equivalents of the yeast cell wall integrity (CWI) sensors. Finally, we discuss recent findings showing an increasing role for CWI signaling in plant immunity and the adaptation to changes in the ionic environment of plant cells. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Mechanochemical Polarization of Contiguous Cell Walls Shapes Plant Pavement Cells.

PubMed

Majda, Mateusz; Grones, Peter; Sintorn, Ida-Maria; Vain, Thomas; Milani, Pascale; Krupinski, Pawel; Zagórska-Marek, Beata; Viotti, Corrado; Jönsson, Henrik; Mellerowicz, Ewa J; Hamant, Olivier; Robert, Stéphanie

2017-11-06

The epidermis of aerial plant organs is thought to be limiting for growth, because it acts as a continuous load-bearing layer, resisting tension. Leaf epidermis contains jigsaw puzzle piece-shaped pavement cells whose shape has been proposed to be a result of subcellular variations in expansion rate that induce local buckling events. Paradoxically, such local compressive buckling should not occur given the tensile stresses across the epidermis. Using computational modeling, we show that the simplest scenario to explain pavement cell shapes within an epidermis under tension must involve mechanical wall heterogeneities across and along the anticlinal pavement cell walls between adjacent cells. Combining genetics, atomic force microscopy, and immunolabeling, we demonstrate that contiguous cell walls indeed exhibit hybrid mechanochemical properties. Such biochemical wall heterogeneities precede wall bending. Altogether, this provides a possible mechanism for the generation of complex plant cell shapes. Copyright © 2017 Elsevier Inc. All rights reserved.

Wall extensibility: its nature, measurement and relationship to plant cell growth

NASA Technical Reports Server (NTRS)

Cosgrove, D. J.

1993-01-01

Expansive growth of plant cells is controlled principally by processes that loosen the wall and enable it to expand irreversibly. The central role of wall relaxation for cell expansion is reviewed. The most common methods for assessing the extension properties of plant cell walls ( wall extensibility') are described, categorized and assessed critically. What emerges are three fundamentally different approaches which test growing cells for their ability (a) to enlarge at different values of turgor, (b) to induce wall relaxation, and (c) to deform elastically or plastically in response to an applied tensile force. Analogous methods with isolated walls are similarly reviewed. The results of these different assays are related to the nature of plant cell growth and pertinent biophysical theory. I argue that the extensibilities' measured by these assays are fundamentally different from one another and that some are more pertinent to growth than others.

Fractal and spectroscopic analysis of soot from internal combustion engines

NASA Astrophysics Data System (ADS)

Swapna, M. S.; Saritha Devi, H. V.; Raj, Vimal; Sankararaman, S.

2018-03-01

Today diesel engines are used worldwide for various applications and very importantly in transportation. Hydrocarbons are the most widespread precursors among carbon sources employed in the production of carbon nanotubes (CNTs). The aging of internal combustion engine is an important parameter in deciding the carbon emission and particulate matter due to incomplete combustion of fuel. In the present work, an attempt has been made for the effective utilization of the aged engines for potential applicationapplications in fuel cells and nanoelectronics. To analyze the impact of aging, the particulate matter rich in carbon content areis collected from diesel engines of different ages. The soot with CNTs is purified by the liquid phase oxidation method and analyzed by Field Emission Scanning Electron Microscopy, High-Resolution Transmission Electron Microscopy, Energy Dispersive Spectroscopy, UV-Visible spectroscopy, Raman spectroscopy and Thermogravimetric analysis. The SEM image contains self-similar patterns probing fractal analysis. The fractal dimensions of the samples are determined by the box counting method. We could find a greater amount of single-walled carbon nanotubes (SWCNTs) in the particulate matter emitted by aged diesel engines and thereby giving information about the combustion efficiency of the engine. The SWCNT rich sample finds a wide range of applicationapplications in nanoelectronics and thereby pointing a potential use of these aged engines.

Generation of hydroxyl radical in isolated pea root cell wall, and the role of cell wall-bound peroxidase, Mn-SOD and phenolics in their production.

PubMed

Kukavica, Biljana; Mojovic, Milos; Vuccinic, Zeljko; Maksimovic, Vuk; Takahama, Umeo; Jovanovic, Sonja Veljovic

2009-02-01

The hydroxyl radical produced in the apoplast has been demonstrated to facilitate cell wall loosening during cell elongation. Cell wall-bound peroxidases (PODs) have been implicated in hydroxyl radical formation. For this mechanism, the apoplast or cell walls should contain the electron donors for (i) H(2)O(2) formation from dioxygen; and (ii) the POD-catalyzed reduction of H(2)O(2) to the hydroxyl radical. The aim of the work was to identify the electron donors in these reactions. In this report, hydroxyl radical (.OH) generation in the cell wall isolated from pea roots was detected in the absence of any exogenous reductants, suggesting that the plant cell wall possesses the capacity to generate .OH in situ. Distinct POD and Mn-superoxide dismutase (Mn-SOD) isoforms different from other cellular isoforms were shown by native gel electropho-resis to be preferably bound to the cell walls. Electron paramagnetic resonance (EPR) spectroscopy of cell wall isolates containing the spin-trapping reagent, 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO), was used for detection of and differentiation between .OH and the superoxide radical (O(2)(-).). The data obtained using POD inhibitors confirmed that tightly bound cell wall PODs are involved in DEPMPO/OH adduct formation. A decrease in DEPMPO/OH adduct formation in the presence of H(2)O(2) scavengers demonstrated that this hydroxyl radical was derived from H(2)O(2). During the generation of .OH, the concentration of quinhydrone structures (as detected by EPR spectroscopy) increased, suggesting that the H(2)O(2) required for the formation of .OH in isolated cell walls is produced during the reduction of O(2) by hydroxycinnamic acids. Cell wall isolates in which the proteins have been denaturated (including the endogenous POD and SOD) did not produce .OH. Addition of exogenous H(2)O(2) again induced the production of .OH, and these were shown to originate from the Fenton reaction with tightly bound metal ions. However, the appearance of the DEPMPO/OOH adduct could also be observed, due to the production of O(2)(-). when endogenous SOD has been inactivated. Also, O(2)(-). was converted to .OH in an in vitro horseradish peroxidase (HRP)/H(2)O(2) system to which exogenous SOD has been added. Taken together with the discovery of the cell wall-bound Mn-SOD isoform, these results support the role of such a cell wall-bound SOD in the formation of .OH jointly with the cell wall-bound POD. According to the above findings, it seems that the hydroxycinnamic acids from the cell wall, acting as reductants, contribute to the formation of H(2)O(2) in the presence of O(2) in an autocatalytic manner, and that POD and Mn-SOD coupled together generate .OH from such H(2)O(2).

A Glycosylphosphatidylinositol Anchor Is Required for Membrane Localization but Dispensable for Cell Wall Association of Chitin Deacetylase 2 in Cryptococcus neoformans

PubMed Central

Gilbert, Nicole M.; Baker, Lorina G.; Specht, Charles A.; Lodge, Jennifer K.

2012-01-01

ABSTRACT Cell wall proteins (CWPs) mediate important cellular processes in fungi, including adhesion, invasion, biofilm formation, and flocculation. The current model of fungal cell wall organization includes a major class of CWPs covalently bound to β-1,6-glucan via a remnant of a glycosylphosphatidylinositol (GPI) anchor. This model was established by studies of ascomycetes more than a decade ago, and relatively little work has been done with other fungi, although the presumption has been that proteins identified in the cell wall which contain a predicted GPI anchor are covalently linked to cell wall glucans. The pathogenic basidiomycete Cryptococcus neoformans encodes >50 putatively GPI-anchored proteins, some of which have been identified in the cell wall. One of these proteins is chitin deacetylase 2 (Cda2), an enzyme responsible for converting chitin to chitosan, a cell wall polymer recently established as a virulence factor for C. neoformans infection of mammalian hosts. Using a combination of biochemistry, molecular biology, and genetics, we show that Cda2 is GPI anchored to membranes but noncovalently associated with the cell wall by means independent of both its GPI anchor and β-1,6-glucan. We also show that Cda2 produces chitosan when localized to the plasma membrane, but association with the cell wall is not essential for this process, thereby providing insight into the mechanism of chitosan biosynthesis. These results increase our understanding of the surface of C. neoformans and provide models of cell walls likely applicable to other undercharacterized basidiomycete pathogenic fungi. PMID:22354955

The mechanics of surface expansion anisotropy in Medicago truncatula root hairs.

PubMed

Dumais, Jacques; Long, Sharon R; Shaw, Sidney L

2004-10-01

Wall expansion in tip-growing cells shows variations according to position and direction. In Medicago truncatula root hairs, wall expansion exhibits a strong meridional gradient with a maximum near the pole of the cell. Root hair cells also show a striking expansion anisotropy, i.e. over most of the dome surface the rate of circumferential wall expansion exceeds the rate of meridional expansion. Concomitant measurements of expansion rates and wall stresses reveal that the extensibility of the cell wall must vary abruptly along the meridian of the cell to maintain the gradient of wall expansion. To determine the mechanical basis of expansion anisotropy, we compared measurements of wall expansion with expansion patterns predicted from wall structural models that were either fully isotropic, transversely isotropic, or fully anisotropic. Our results indicate that a model based on a transversely isotropic wall structure can provide a good fit of the data although a fully anisotropic model offers the best fit overall. We discuss how such mechanical properties could be controlled at the microstructural level.

Functional duality of the cell wall.

PubMed

Latgé, Jean-Paul; Beauvais, Anne

2014-08-01

The polysaccharide cell wall is the extracellular armour of the fungal cell. Although essential in the protection of the fungal cell against aggressive external stresses, the biosynthesis of the polysaccharide core is poorly understood. For a long time it was considered that this cell wall skeleton was a fixed structure whose role was only to be sensed as non-self by the host and consequently trigger the defence response. It is now known that the cell wall polysaccharide composition and localization continuously change to adapt to their environment and that these modifications help the fungus to escape from the immune system. Moreover, cell wall polysaccharides could function as true virulence factors. Copyright © 2014 Elsevier Ltd. All rights reserved.

Distinct Cell Wall Architectures in Seed Endosperms in Representatives of the Brassicaceae and Solanaceae1[C][W][OA

PubMed Central

Lee, Kieran J.D.; Dekkers, Bas J.W.; Steinbrecher, Tina; Walsh, Cherie T.; Bacic, Antony; Bentsink, Leónie; Leubner-Metzger, Gerhard; Knox, J. Paul

2012-01-01

In some species, a crucial role has been demonstrated for the seed endosperm during germination. The endosperm has been shown to integrate environmental cues with hormonal networks that underpin dormancy and seed germination, a process that involves the action of cell wall remodeling enzymes (CWREs). Here, we examine the cell wall architectures of the endosperms of two related Brassicaceae, Arabidopsis (Arabidopsis thaliana) and the close relative Lepidium (Lepidium sativum), and that of the Solanaceous species, tobacco (Nicotiana tabacum). The Brassicaceae species have a similar cell wall architecture that is rich in pectic homogalacturonan, arabinan, and xyloglucan. Distinctive features of the tobacco endosperm that are absent in the Brassicaceae representatives are major tissue asymmetries in cell wall structural components that reflect the future site of radicle emergence and abundant heteromannan. Cell wall architecture of the micropylar endosperm of tobacco seeds has structural components similar to those seen in Arabidopsis and Lepidium endosperms. In situ and biomechanical analyses were used to study changes in endosperms during seed germination and suggest a role for mannan degradation in tobacco. In the case of the Brassicaceae representatives, the structurally homogeneous cell walls of the endosperm can be acted on by spatially regulated CWRE expression. Genetic manipulations of cell wall components present in the Arabidopsis seed endosperm demonstrate the impact of cell wall architectural changes on germination kinetics. PMID:22961130

An unusual xylan in Arabidopsis primary cell walls is synthesised by GUX3, IRX9L, IRX10L and IRX14

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mortimer, Jenny C.; Faria-Blanc, Nuno; Yu, Xiaolan

Xylan is a crucial component of many plant primary and secondary cell walls. However, the structure and function of xylan in the dicotyledon primary cell wall is not well understood. Here, we characterized a xylan that is specific to tissues enriched in Arabidopsis primary cell walls. Unlike previously described xylans, this xylan carries a pentose linked 1–2 to the α-1,2-d-glucuronic acid (GlcA) side chains on the β-1,4-Xyl backbone. The frequent and precisely regular spacing of GlcA substitutions every six xylosyl residues along the backbone is also unlike that previously observed in secondary cell wall xylan. Molecular genetics, in vitro assays,more » and expression data suggest that IRX9L, IRX10L and IRX14 are required for xylan backbone synthesis in primary cell wall synthesising tissues. IRX9 and IRX10 are not involved in the primary cell wall xylan synthesis but are functionally exchangeable with IRX9L and IRX10L. GUX3 is the only glucuronyltransferase required for the addition of the GlcA decorations on the xylan. Lastly, the differences in xylan structure in primary versus secondary cell walls might reflect the different roles in cross-linking and interaction with other cell wall components.« less

An unusual xylan in Arabidopsis primary cell walls is synthesised by GUX3, IRX9L, IRX10L and IRX14

DOE PAGES

Mortimer, Jenny C.; Faria-Blanc, Nuno; Yu, Xiaolan; ...

2015-06-04

Xylan is a crucial component of many plant primary and secondary cell walls. However, the structure and function of xylan in the dicotyledon primary cell wall is not well understood. Here, we characterized a xylan that is specific to tissues enriched in Arabidopsis primary cell walls. Unlike previously described xylans, this xylan carries a pentose linked 1–2 to the α-1,2-d-glucuronic acid (GlcA) side chains on the β-1,4-Xyl backbone. The frequent and precisely regular spacing of GlcA substitutions every six xylosyl residues along the backbone is also unlike that previously observed in secondary cell wall xylan. Molecular genetics, in vitro assays,more » and expression data suggest that IRX9L, IRX10L and IRX14 are required for xylan backbone synthesis in primary cell wall synthesising tissues. IRX9 and IRX10 are not involved in the primary cell wall xylan synthesis but are functionally exchangeable with IRX9L and IRX10L. GUX3 is the only glucuronyltransferase required for the addition of the GlcA decorations on the xylan. Lastly, the differences in xylan structure in primary versus secondary cell walls might reflect the different roles in cross-linking and interaction with other cell wall components.« less

Spatially and temporally restricted expression of PtrMYB021 regulates secondary cell wall formation in Arabidopsis

DOE PAGES

Wang, Wei; Li, Eryang; Porth, Ilga; ...

2016-02-02

Among the R2R3 MYB transcription factors that involve in the regulation of secondary cell wall formation in Arabidopsis, MYB46 alone is sufficient to induce the entire secondary cell wall biosynthesis program. PtrMYB021, the poplar homolog of MYB46, has been reported to regulate secondary cell wall formation when expressed in Arabidopsis. We report here that spatially and temporally restricted expression of PtrMYB021 is critical for its function in regulating secondary cell wall formation. By using quantitative RT-PCR, we found that PtrMYB021 was expressed primarily in xylem tissues. When expressed in Arabidopsis under the control of PtrCesA8, but not the 35S promoter,more » PtrMYB021 increased secondary cell wall thickness, which is likely caused by increased lignification as well as changes in cell wall carbohydrate composition. Consistent with this, elevated expression of lignin and cellulose biosynthetic genes were observed in the transgenic plants. Finally, when expressed in Arabidopsis protoplasts as fusion proteins to the Gal4 DNA binding domain, PtrMYB021 activated the reporter gene Gal4-GUS. In summary, our results suggest that PtrMYB021 is a transcriptional activator, and spatially and temporally restricted expression of PtrMYB021 in Arabidopsis regulates secondary cell wall formation by activating a subset of secondary cell wall biosynthesis genes.« less

Spatially and temporally restricted expression of PtrMYB021 regulates secondary cell wall formation in Arabidopsis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Li, Eryang; Porth, Ilga

Among the R2R3 MYB transcription factors that involve in the regulation of secondary cell wall formation in Arabidopsis, MYB46 alone is sufficient to induce the entire secondary cell wall biosynthesis program. PtrMYB021, the poplar homolog of MYB46, has been reported to regulate secondary cell wall formation when expressed in Arabidopsis. We report here that spatially and temporally restricted expression of PtrMYB021 is critical for its function in regulating secondary cell wall formation. By using quantitative RT-PCR, we found that PtrMYB021 was expressed primarily in xylem tissues. When expressed in Arabidopsis under the control of PtrCesA8, but not the 35S promoter,more » PtrMYB021 increased secondary cell wall thickness, which is likely caused by increased lignification as well as changes in cell wall carbohydrate composition. Consistent with this, elevated expression of lignin and cellulose biosynthetic genes were observed in the transgenic plants. Finally, when expressed in Arabidopsis protoplasts as fusion proteins to the Gal4 DNA binding domain, PtrMYB021 activated the reporter gene Gal4-GUS. In summary, our results suggest that PtrMYB021 is a transcriptional activator, and spatially and temporally restricted expression of PtrMYB021 in Arabidopsis regulates secondary cell wall formation by activating a subset of secondary cell wall biosynthesis genes.« less

Forage digestibility: the intersection of cell wall lignification and plant tissue anatomy

USDA-ARS?s Scientific Manuscript database

Cellulose and the other polysaccharides present in forage cell walls can be completely degraded by the rumen microflora but only when these polysaccharides have been isolated from the wall and all matrix structures eliminated. Understanding how cell wall component interactions limit microbial degrad...

A synthetic leaf: the biomimetic potential of graphene oxide

NASA Astrophysics Data System (ADS)

Lamb, Marilla; Koch, George W.; Morgan, Eric R.; Shafer, Michael W.

2015-03-01

Emerging materials such as graphene oxide (GO) have micro and nano features that are functionally similar to those in plant cell walls involved in water transport. Therefore, it may now be possible to design and build biomimetic trees to lift water via mechanisms similar to those employed by trees, allowing for potential applications such as passive water pumping, filtering, and evaporative cooling. The tallest trees can raise large volumes of water to over 100 meters using only the vapor pressure gradient between their leaves and the atmosphere. This phenomenon occurs in all terrestrial plants when capillary forces generated in the microscopic pores in the cell walls of leaves are collectively applied to large diameter xylem conduits. The design of a synthetic tree that mimics these mechanisms will allow water to be moved to heights greater than is currently possible by any engineered system that does not require the use of a positive pressure pump. We are testing the suitability of membranous GO as the leaf of a synthetic tree and present an analysis in support of this design. In addition, we include results from a preliminary design using ceramics.

Chalcone Synthase (CHS) Gene Suppression in Flax Leads to Changes in Wall Synthesis and Sensing Genes, Cell Wall Chemistry and Stem Morphology Parameters

PubMed Central

Zuk, Magdalena; Działo, Magdalena; Richter, Dorota; Dymińska, Lucyna; Matuła, Jan; Kotecki, Andrzej; Hanuza, Jerzy; Szopa, Jan

2016-01-01

The chalcone synthase (CHS) gene controls the first step in the flavonoid biosynthesis. In flax, CHS down-regulation resulted in tannin accumulation and reduction in lignin synthesis, but plant growth was not affected. This suggests that lignin content and thus cell wall characteristics might be modulated through CHS activity. This study investigated the possibility that CHS affects cell wall sensing as well as polymer content and arrangement. CHS-suppressed and thus lignin-reduced plants showed significant changes in expression of genes involved in both synthesis of components and cell wall sensing. This was accompanied by increased levels of cellulose and hemicellulose. CHS-reduced flax also showed significant changes in morphology and arrangement of the cell wall. The stem tissue layers were enlarged averagely twofold compared to the control, and the number of fiber cells more than doubled. The stem morphology changes were accompanied by reduction of the crystallinity index of the cell wall. CHS silencing induces a signal transduction cascade that leads to modification of plant metabolism in a wide range and thus cell wall structure. PMID:27446124

If walls could talk

NASA Technical Reports Server (NTRS)

Braam, J.; McIntire, L. V. (Principal Investigator)

1999-01-01

The plant cell wall is very complex, both in structure and function. The wall components and the mechanical properties of the wall have been implicated in conveying information that is important for morphogenesis. Proteoglycans, fragments of polysaccharides and the structural integrity of the wall may relay signals that influence cellular differentiation and growth control. Furthering our knowledge of cell wall structure and function is likely to have a profound impact on our understanding of how plant cells communicate with the extracellular environment.

Arrangement of Cellulose Microfibrils in Walls of Elongating Parenchyma Cells

PubMed Central

Setterfield, G.; Bayley, S. T.

1958-01-01

The arrangement of cellulose microfibrils in walls of elongating parenchyma cells of Avena coleoptiles, onion roots, and celery petioles was studied in polarizing and electron microscopes by examining whole cell walls and sections. Walls of these cells consist firstly of regions containing the primary pit fields and composed of microfibrils oriented predominantly transversely. The transverse microfibrils show a progressive disorientation from the inside to the outside of the wall which is consistent with the multinet model of wall growth. Between the pit-field regions and running the length of the cells are ribs composed of longitudinally oriented microfibrils. Two types of rib have been found at all stages of cell elongation. In some regions, the wall appears to consist entirely of longitudinal microfibrils so that the rib forms an integral part of the wall. At the edges of such ribs the microfibrils can be seen to change direction from longitudinal in the rib to transverse in the pit-field region. Often, however, the rib appears to consist of an extra separate layer of longitudinal microfibrils outside a continuous wall of transverse microfibrils. These ribs are quite distinct from secondary wall, which consists of longitudinal microfibrils deposited within the primary wall after elongation has ceased. It is evident that the arrangement of cellulose microfibrils in a primary wall can be complex and is probably an expression of specific cellular differentiation. PMID:13563544

Effect of Inhibition of Deoxyribonucleic Acid and Protein Synthesis on the Direction of Cell Wall Growth in Streptococcus faecalis

PubMed Central

Higgins, M. L.; Daneo-Moore, L.; Boothby, D.; Shockman, G. D.

1974-01-01

Selective inhibition of protein synthesis in Streptococcus faecalis (ATCC 9790) was accompanied by a rapid and severe inhibition of cell division and a reduction of enlargement of cellular surface area. Continued synthesis of cell wall polymers resulted in rapid thickening of the wall to an extent not seen in exponential-phase populations. Thus, the normal direction of wall growth was changed from a preferential feeding out of new wall surface to that of thickening existing cell surfaces. However, the overall manner in which the wall thickened, from nascent septa toward polar regions, was the same in both exponential-phase and inhibited populations. In contrast, selective inhibition of deoxyribonucleic acid (DNA) synthesis using mitomycin C was accompanied by an increase in cellular surface area and by division of about 80% of the cells in random populations. Little or no wall thickening was observed until the synthesis of macromolecules other than DNA was impaired and further cell division ceased. Concomitant inhibition of both DNA and protein synthesis inhibited cell division but permitted an increase in average cell volume. In such doubly inhibited cells, walls thickened less than in cells inhibited for protein synthesis only. On the basis of the results obtained, a model for cell surface enlargement and cell division is presented. The model proposes that: (i) each wall enlargement site is influenced by an individual chromosome replication cycle; (ii) during chromosome replication peripheral surface enlargement would be favored over thickening (or septation); (iii) a signal associated with chromosome termination would favor thickening (and septation) at the expense of surface enlargement; and (iv) a factor or signal related to protein synthesis would be required for one or more of the near terminal stages of cell division or cell separation, or both. Images PMID:4133352

Transient Temperature Analysis in a System of Thin Shells Combined with Convective and Radiative Cooling

NASA Astrophysics Data System (ADS)

Prasad, Ravindra; Samria, N. K.

1989-01-01

The problem considered has applications in the transient thermal analysis and time for attaining the steady state of the cylinder wall and cylinder head of an air-cooled internal-combustion engine. Numerical calculations based on finite difference approximations are carried out to assess the thermal response in a system of thin cylindrical and spherical shells having hot gases inside with convective boundary conditions. The outside surface is exposed to cooling medium where it looses heat by natural convection and radiation. As a special case, when radius is large, the surface may be considered to be a plane wall. The cylinder cover and cylinder wall of an internal-combustion engine are considered to be a plane wall for a comparatively higher ratio of cylinder diameter to the thickness of the wall, i.e., whend/δ varies from 80 to 100. A plot of temperature-time history and heat flow rates have been obtained.

Insights into cell wall structure of Sida hermaphrodita and its influence on recalcitrance.

PubMed

Damm, Tatjana; Pattathil, Sivakumar; Günl, Markus; Jablonowski, Nicolai David; O'Neill, Malcolm; Grün, Katharina Susanne; Grande, Philipp Michael; Leitner, Walter; Schurr, Ulrich; Usadel, Björn; Klose, Holger

2017-07-15

The perennial plant Sida hermaphrodita (Sida) is attracting attention as potential energy crop. Here, the first detailed view on non-cellulosic Sida cell wall polysaccharide composition, structure and architecture is given. Cell walls were prepared from Sida stems and sequentially extracted with aqueous buffers and alkali. The structures of the quantitatively predominant polysaccharides present in each fraction were determined by biochemical characterization, glycome profiling and mass spectrometry. The amounts of glucose released by Accellerase-1500 ® treatment of the cell wall and the cell wall residue remaining after each extraction were used to assess the roles of pectin and hemicellulose in the recalcitrance of Sida biomass. 4-O-Methyl glucuronoxylan with a low proportion of side substitutions was identified as the major non-cellulosic glycan component of Sida stem cell walls. Pectic polysaccharides and xylans were found to be associated with lignin, suggesting that these polysaccharides have roles in Sida cell wall recalcitrance to enzymatic hydrolysis. Copyright © 2017 Elsevier Ltd. All rights reserved.

Application of X-ray and neutron small angle scattering techniques to study the hierarchical structure of plant cell walls: a review.

PubMed

Martínez-Sanz, Marta; Gidley, Michael J; Gilbert, Elliot P

2015-07-10

Plant cell walls present an extremely complex structure of hierarchically assembled cellulose microfibrils embedded in a multi-component matrix. The biosynthesis process determines the mechanism of cellulose crystallisation and assembly, as well as the interaction of cellulose with other cell wall components. Thus, a knowledge of cellulose microfibril and bundle architecture, and the structural role of matrix components, is crucial for understanding cell wall functional and technological roles. Small angle scattering techniques, combined with complementary methods, provide an efficient approach to characterise plant cell walls, covering a broad and relevant size range while minimising experimental artefacts derived from sample treatment. Given the system complexity, approaches such as component extraction and the use of plant cell wall analogues are typically employed to enable the interpretation of experimental results. This review summarises the current research status on the characterisation of the hierarchical structure of plant cell walls using small angle scattering techniques. Crown Copyright © 2015. Published by Elsevier Ltd. All rights reserved.

Coolant Design System for Liquid Propellant Aerospike Engines

NASA Astrophysics Data System (ADS)

McConnell, Miranda; Branam, Richard

2015-11-01

Liquid propellant rocket engines burn at incredibly high temperatures making it difficult to design an effective coolant system. These particular engines prove to be extremely useful by powering the rocket with a variable thrust that is ideal for space travel. When combined with aerospike engine nozzles, which provide maximum thrust efficiency, this class of rockets offers a promising future for rocketry. In order to troubleshoot the problems that high combustion chamber temperatures pose, this research took a computational approach to heat analysis. Chambers milled into the combustion chamber walls, lined by a copper cover, were tested for their efficiency in cooling the hot copper wall. Various aspect ratios and coolants were explored for the maximum wall temperature by developing our own MATLAB code. The code uses a nodal temperature analysis with conduction and convection equations and assumes no internal heat generation. This heat transfer research will show oxygen is a better coolant than water, and higher aspect ratios are less efficient at cooling. This project funded by NSF REU Grant 1358991.

Analyzing Cell Wall Elasticity After Hormone Treatment: An Example Using Tobacco BY-2 Cells and Auxin.

PubMed

Braybrook, Siobhan A

2017-01-01

Atomic force microscopy, and related nano-indentation techniques, is a valuable tool for analyzing the elastic properties of plant cell walls as they relate to changes in cell wall chemistry, changes in development, and response to hormones. Within this chapter I will describe a method for analyzing the effect of the phytohormone auxin on the cell wall elasticity of tobacco BY-2 cells. This general method may be easily altered for different experimental systems and hormones of interest.

Ectopic lignification in primary cellulose-deficient cell walls of maize cell suspension cultures.

PubMed

Mélida, Hugo; Largo-Gosens, Asier; Novo-Uzal, Esther; Santiago, Rogelio; Pomar, Federico; García, Pedro; García-Angulo, Penélope; Acebes, José Luis; Álvarez, Jesús; Encina, Antonio

2015-04-01

Maize (Zea mays L.) suspension-cultured cells with up to 70% less cellulose were obtained by stepwise habituation to dichlobenil (DCB), a cellulose biosynthesis inhibitor. Cellulose deficiency was accompanied by marked changes in cell wall matrix polysaccharides and phenolics as revealed by Fourier transform infrared (FTIR) spectroscopy. Cell wall compositional analysis indicated that the cellulose-deficient cell walls showed an enhancement of highly branched and cross-linked arabinoxylans, as well as an increased content in ferulic acid, diferulates and p-coumaric acid, and the presence of a polymer that stained positive for phloroglucinol. In accordance with this, cellulose-deficient cell walls showed a fivefold increase in Klason-type lignin. Thioacidolysis/GC-MS analysis of cellulose-deficient cell walls indicated the presence of a lignin-like polymer with a Syringyl/Guaiacyl ratio of 1.45, which differed from the sensu stricto stress-related lignin that arose in response to short-term DCB-treatments. Gene expression analysis of these cells indicated an overexpression of genes specific for the biosynthesis of monolignol units of lignin. A study of stress signaling pathways revealed an overexpression of some of the jasmonate signaling pathway genes, which might trigger ectopic lignification in response to cell wall integrity disruptions. In summary, the structural plasticity of primary cell walls is proven, since a lignification process is possible in response to cellulose impoverishment. © 2015 Institute of Botany, Chinese Academy of Sciences.

Non-lignified helical cell wall thickenings in root cortical cells of Aspleniaceae (Polypodiales): histology and taxonomical significance

PubMed Central

Leroux, O.; Bagniewska-Zadworna, A.; Rambe, S. K.; Knox, J. P.; Marcus, S. E.; Bellefroid, E.; Stubbe, D.; Chabbert, B.; Habrant, A.; Claeys, M.; Viane, R. L. L.

2011-01-01

Background and Aims Extraxylary helical cell wall thickenings in vascular plants are not well documented, except for those in orchid velamen tissues which have been studied extensively. Reports on their occurrence in ferns exist, but detailed information is missing. The aim of this study is to focus on the broad patterns of structure and composition and to study the taxonomic occurrence of helical cell wall thickenings in the fern family Aspleniaceae. Methods Structural and compositional aspects of roots have been examined by means of light, electron, epifluorescence and laser scanning confocal microscopy. To assess the taxonomical distribution of helical cell wall thickenings a molecular phylogenetic analysis based on rbcL sequences of 64 taxa was performed. Key Results The helical cell wall thickenings of all examined species showed considerable uniformity of design. The pattern consists of helical, regularly bifurcating and anastomosing strands. Compositionally, the cell wall thickenings were found to be rich in homogalacturonan, cellulose, mannan and xyloglucan. Thioacidolysis confirmed our negative phloroglucinol staining tests, demonstrating the absence of lignins in the root cortex. All taxa with helical cell wall thickenings formed a monophyletic group supported by a 100 % bootstrap value and composed of mainly epiphytic species. Conclusions This is the first report of non-lignified pectin-rich secondary cell walls in ferns. Based on our molecular analysis, we reject the hypothesis of parallel evolution of helical cell wall thickenings in Aspleniaceae. Helical cell wall thickenings can mechanically stabilize the cortex tissue, allowing maximal uptake of water and nutrients during rainfall events. In addition, it can also act as a boundary layer increasing the diffusive pathway towards the atmosphere, preventing desiccation of the stele of epiphytic growing species. PMID:21118842

Anatomical structure overrides temperature controls on magnesium uptake - calcification in the Arctic/subarctic coralline algae Leptophytum laeve and Kvaleya epilaeve (Rhodophyta; Corallinales)

NASA Astrophysics Data System (ADS)

Nash, Merinda C.; Adey, Walter

2018-02-01

Calcified coralline red algae are ecologically key organisms in photic benthic environments. In recent decades they have become important climate proxies, especially in the Arctic and subarctic. It has been widely accepted that magnesium content in coralline tissues is directly a function of ambient temperature, and this is a primary basis for their value as a climate archive. In this paper we show for two genera of Arctic/subarctic corallines, Leptophytum laeve and Kvaleya epilaeve, that previously unrecognised complex tissue and cell wall anatomy bears a variety of basal signatures for Mg content, with the accepted temperature relationship being secondary. The interfilament carbonate has lower Mg than adjacent cell walls and the hypothallial cell walls have the highest Mg content. The internal structure of the hypothallial cell walls can differ substantially from the perithallial radial cell wall structure. Using high-magnification scanning electron microscopy and etching we expose the nanometre-scale structures within the cell walls and interfilament. Fibrils concentrate at the internal and external edges of the cell walls. Fibrils ˜ 10 nm thick appear to thread through the radial Mg-calcite grains and form concentric bands within the cell wall. This banding may control Mg distribution within the cell. Similar fibril banding is present in the hypothallial cell walls but not the interfilament. Climate archiving with corallines can achieve greater precision with recognition of these parameters.

Plant cell walls throughout evolution: towards a molecular understanding of their design principles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sarkar, Purbasha; Bosneaga, Elena; Auer, Manfred

Throughout their life, plants typically remain in one location utilizing sunlight for the synthesis of carbohydrates, which serve as their sole source of energy as well as building blocks of a protective extracellular matrix, called the cell wall. During the course of evolution, plants have repeatedly adapted to their respective niche,which is reflected in the changes of their body plan and the specific design of cell walls. Cell walls not only changed throughout evolution but also are constantly remodelled and reconstructed during the development of an individual plant, and in response to environmental stress or pathogen attacks. Carbohydrate-rich cell wallsmore » display complex designs, which together with the presence of phenolic polymers constitutes a barrier for microbes, fungi, and animals. Throughout evolution microbes have co-evolved strategies for efficient breakdown of cell walls. Our current understanding of cell walls and their evolutionary changes are limited as our knowledge is mainly derived from biochemical and genetic studies, complemented by a few targeted yet very informative imaging studies. Comprehensive plant cell wall models will aid in the re-design of plant cell walls for the purpose of commercially viable lignocellulosic biofuel production as well as for the timber, textile, and paper industries. Such knowledge will also be of great interest in the context of agriculture and to plant biologists in general. It is expected that detailed plant cell wall models will require integrated correlative multimodal, multiscale imaging and modelling approaches, which are currently underway.« less

Endomembrane proteomics reveals putative enzymes involved in cell wall metabolism in wheat grain outer layers

PubMed Central

Chateigner-Boutin, Anne-Laure; Suliman, Muhtadi; Bouchet, Brigitte; Alvarado, Camille; Lollier, Virginie; Rogniaux, Hélène; Guillon, Fabienne; Larré, Colette

2015-01-01

Cereal grain outer layers fulfil essential functions for the developing seed such as supplying energy and providing protection. In the food industry, the grain outer layers called ‘the bran’ is valuable since it is rich in dietary fibre and other beneficial nutriments. The outer layers comprise several tissues with a high content in cell wall material. The cell wall composition of the grain peripheral tissues was investigated with specific probes at a stage of active cell wall synthesis. Considerable wall diversity between cell types was revealed. To identify the cellular machinery involved in cell wall synthesis, a subcellular proteomic approach was used targeting the Golgi apparatus where most cell wall polysaccharides are synthesized. The tissues were dissected into outer pericarp and intermediate layers where 822 and 1304 proteins were identified respectively. Many carbohydrate-active enzymes were revealed: some in the two peripheral grain fractions, others only in one tissue. Several protein families specific to one fraction and with characterized homologs in other species might be related to the specific detection of a polysaccharide in a particular cell layer. This report provides new information on grain cell walls and its biosynthesis in the valuable outer tissues, which are poorly studied so far. A better understanding of the mechanisms controlling cell wall composition could help to improve several quality traits of cereal products (e.g. dietary fibre content, biomass conversion to biofuel). PMID:25769308

Characterization of a Lignified Secondary Phloem Fibre‐deficient Mutant of Jute (Corchorus capsularis)

PubMed Central

SENGUPTA, GARGI; PALIT, P.

2004-01-01

• Background and Aims High lignin content of lignocellulose jute fibre does not favour its utilization in making finer fabrics and other value‐added products. To aid the development of low‐lignin jute fibre, this study aimed to identify a phloem fibre mutant with reduced lignin. • Methods An x‐ray‐induced mutant line (CMU) of jute (Corchorus capsularis) was morphologically evaluated and the accession (CMU 013) with the most undulated phenotype was compared with its normal parent (JRC 212) for its growth, secondary fibre development and lignification of the fibre cell wall. • Key Results The normal and mutant plants showed similar leaf photosynthetic rates. The mutant grew more slowly, had shorter internodes and yielded much less fibre after retting. The fibre of the mutant contained 50 % less lignin but comparatively more cellulose than that of the normal type. Differentiation of primary and secondary vascular tissues throughout the CMU 013 stem was regular but it did not have secondary phloem fibre bundles as in JRC 212. Instead, a few thin‐walled, less lignified fibre cells formed uni‐ or biseriate radial rows within the phloem wedges of the middle stem. The lower and earliest developed part of the mutant stem had no lignified fibre cells. This developmental deficiency in lignification of fibre cells was correlated to a similar deficiency in phenylalanine ammonia lyase activity, but not peroxidase activity, in the bark tissue along the stem axis. In spite of severe reduction in lignin synthesis in the phloem cells this mutant functioned normally and bred true. • Conclusions In view of the observations made, the mutant is designated as deficient lignified phloem fibre (dlpf). This mutant may be utilized to engineer low‐lignin jute fibre strains and may also serve as a model to study the positional information that coordinates secondary wall thickening of fibre cells. PMID:14707004

Bioinspired metal-cell wall-metal sandwich structure on an individual bacterial cell scaffold.

PubMed

Zhang, Xiaoliang; Yu, Mei; Liu, Jianhua; Li, Songmei

2012-08-25

Pd nanoparticles were introduced to individual Bacillus cells and dispersedly anchored on both the inside and outside of the cell walls. The anchored nanoparticles served as "seeds" to drive the formation of double metallic layers forming a metal-cell wall-metal sandwich structure at the single-cell level.

Aquaporin-1 facilitates pressure-driven water flow across the aortic endothelium.

PubMed

Nguyen, Tieuvi; Toussaint, Jimmy; Xue, Yan; Raval, Chirag; Cancel, Limary; Russell, Stewart; Shou, Yixin; Sedes, Omer; Sun, Yu; Yakobov, Roman; Tarbell, John M; Jan, Kung-ming; Rumschitzki, David S

2015-05-01

Aquaporin-1, a ubiquitous water channel membrane protein, is a major contributor to cell membrane osmotic water permeability. Arteries are the physiological system where hydrostatic dominates osmotic pressure differences. In the present study, we show that the walls of large conduit arteries constitute the first example where hydrostatic pressure drives aquaporin-1-mediated transcellular/transendothelial flow. We studied cultured aortic endothelial cell monolayers and excised whole aortas of male Sprague-Dawley rats with intact and inhibited aquaporin-1 activity and with normal and knocked down aquaporin-1 expression. We subjected these systems to transmural hydrostatic pressure differences at zero osmotic pressure differences. Impaired aquaporin-1 endothelia consistently showed reduced engineering flow metrics (transendothelial water flux and hydraulic conductivity). In vitro experiments with tracers that only cross the endothelium paracellularly showed that changes in junctional transport cannot explain these reductions. Percent reductions in whole aortic wall hydraulic conductivity with either chemical blocking or knockdown of aquaporin-1 differed at low and high transmural pressures. This observation highlights how aquaporin-1 expression likely directly influences aortic wall mechanics by changing the critical transmural pressure at which its sparse subendothelial intima compresses. Such compression increases transwall flow resistance. Our endothelial and historic erythrocyte membrane aquaporin density estimates were consistent. In conclusion, aquaporin-1 significantly contributes to hydrostatic pressure-driven water transport across aortic endothelial monolayers, both in culture and in whole rat aortas. This transport, and parallel junctional flow, can dilute solutes that entered the wall paracellularly or through endothelial monolayer disruptions. Lower atherogenic precursor solute concentrations may slow their intimal entrainment kinetics. Copyright © 2015 the American Physiological Society.

Aquaporin-1 facilitates pressure-driven water flow across the aortic endothelium

PubMed Central

Nguyen, Tieuvi; Toussaint, Jimmy; Xue, Yan; Raval, Chirag; Cancel, Limary; Russell, Stewart; Shou, Yixin; Sedes, Omer; Sun, Yu; Yakobov, Roman; Tarbell, John M.; Jan, Kung-ming

2015-01-01

Aquaporin-1, a ubiquitous water channel membrane protein, is a major contributor to cell membrane osmotic water permeability. Arteries are the physiological system where hydrostatic dominates osmotic pressure differences. In the present study, we show that the walls of large conduit arteries constitute the first example where hydrostatic pressure drives aquaporin-1-mediated transcellular/transendothelial flow. We studied cultured aortic endothelial cell monolayers and excised whole aortas of male Sprague-Dawley rats with intact and inhibited aquaporin-1 activity and with normal and knocked down aquaporin-1 expression. We subjected these systems to transmural hydrostatic pressure differences at zero osmotic pressure differences. Impaired aquaporin-1 endothelia consistently showed reduced engineering flow metrics (transendothelial water flux and hydraulic conductivity). In vitro experiments with tracers that only cross the endothelium paracellularly showed that changes in junctional transport cannot explain these reductions. Percent reductions in whole aortic wall hydraulic conductivity with either chemical blocking or knockdown of aquaporin-1 differed at low and high transmural pressures. This observation highlights how aquaporin-1 expression likely directly influences aortic wall mechanics by changing the critical transmural pressure at which its sparse subendothelial intima compresses. Such compression increases transwall flow resistance. Our endothelial and historic erythrocyte membrane aquaporin density estimates were consistent. In conclusion, aquaporin-1 significantly contributes to hydrostatic pressure-driven water transport across aortic endothelial monolayers, both in culture and in whole rat aortas. This transport, and parallel junctional flow, can dilute solutes that entered the wall paracellularly or through endothelial monolayer disruptions. Lower atherogenic precursor solute concentrations may slow their intimal entrainment kinetics. PMID:25659484

Post-Synthetic Defucosylation of AGP by Aspergillus nidulans α-1,2-Fucosidase Expressed in Arabidopsis Apoplast Induces Compensatory Upregulation of α-1,2-Fucosyltransferases.

PubMed

Pogorelko, Gennady V; Reem, Nathan T; Young, Zachary T; Chambers, Lauran; Zabotina, Olga A

2016-01-01

Cell walls are essential components of plant cells which perform a variety of important functions for the different cell types, tissues and organs of a plant. Besides mechanical function providing cell shape, cell walls participate in intercellular communication, defense during plant-microbe interactions, and plant growth. The plant cell wall consists predominantly of polysaccharides with the addition of structural glycoproteins, phenolic esters, minerals, lignin, and associated enzymes. Alterations in the cell wall composition created through either changes in biosynthesis of specific constituents or their post-synthetic modifications in the apoplast compromise cell wall integrity and frequently induce plant compensatory responses as a result of these alterations. Here we report that post-synthetic removal of fucose residues specifically from arabinogalactan proteins in the Arabidopsis plant cell wall induces differential expression of fucosyltransferases and leads to the root and hypocotyl elongation changes. These results demonstrate that the post-synthetic modification of cell wall components presents a valuable approach to investigate the potential signaling pathways induced during plant responses to such modifications that usually occur during plant development and stress responses.

Preparation of Purified Gram-positive Bacterial Cell Wall and Detection in Placenta and Fetal Tissues

PubMed Central

Mann, Beth; Loh, Lip Nam; Gao, Geli; Tuomanen, Elaine

2017-01-01

Cell wall is a complex biopolymer on the surface of all Gram-positive bacteria. During infection, cell wall is recognized by the innate immune receptor Toll-like receptor 2 causing intense inflammation and tissue damage. In animal models, cell wall traffics from the blood stream to many organs in the body, including brain, heart, placenta and fetus. This protocol describes how to prepare purified cell wall from Streptococcus pneumoniae, detect its distribution in animal tissues, and study the tissue response using the placenta and fetal brain as examples. PMID:28573167

Early local differentiation of the cell wall matrix defines the contact sites in lobed mesophyll cells of Zea mays.

PubMed

Giannoutsou, E; Sotiriou, P; Apostolakos, P; Galatis, B

2013-10-01

The morphogenesis of lobed mesophyll cells (MCs) is highly controlled and coupled with intercellular space formation. Cortical microtubule rings define the number and the position of MC isthmi. This work investigated early events of MC morphogenesis, especially the mechanism defining the position of contacts between MCs. The distributions of plasmodesmata, the hemicelluloses callose and (1 → 3,1 → 4)-β-d-glucans (MLGs) and the pectin epitopes recognized by the 2F4, JIM5, JIM7 and LM6 antibodies were studied in the cell walls of Zea mays MCs. Matrix cell wall polysaccharides were immunolocalized in hand-made sections and in sections of material embedded in LR White resin. Callose was also localized using aniline blue in hand-made sections. Plasmodesmata distribution was examined by transmission electron microscopy. Before reorganization of the dispersed cortical microtubules into microtubule rings, particular bands of the longitudinal MC walls, where the MC contacts will form, locally differentiate by selective (1) deposition of callose and the pectin epitopes recognized by the 2F4, LM6, JIM5 and JIM7 antibodies, (2) degradation of MLGs and (3) formation of secondary plasmodesmata clusterings. This cell wall matrix differentiation persists in cell contacts of mature MCs. Simultaneously, the wall bands between those of future cell contacts differentiate with (1) deposition of local cell wall thickenings including cellulose microfibrils, (2) preferential presence of MLGs, (3) absence of callose and (4) transient presence of the pectins identified by the JIM5 and JIM7 antibodies. The wall areas between cell contacts expand determinately to form the cell isthmi and the cell lobes. The morphogenesis of lobed MCs is characterized by the early patterned differentiation of two distinct cell wall subdomains, defining the sites of the future MC contacts and of the future MC isthmi respectively. This patterned cell wall differentiation precedes cortical microtubule reorganization and may define microtubule ring disposition.

Creep behavior of soil nail walls in high plasticity index (PI) soils : technical report.

DOT National Transportation Integrated Search

2017-04-01

An aspect of particular concern in the Geotechnical Engineering Circular No. 7: Soil Nail Walls (i.e., the soil : nail wall manual and construction guidelines) is the creep behavior of soil nail systems in high-plasticity : clays. This research proje...

Stomatal cell wall composition: distinctive structural patterns associated with different phylogenetic groups

PubMed Central

Shtein, Ilana; Shelef, Yaniv; Marom, Ziv; Zelinger, Einat; Schwartz, Amnon; Popper, Zoë A.; Bar-On, Benny

2017-01-01

Background and Aims Stomatal morphology and function have remained largely conserved throughout ∼400 million years of plant evolution. However, plant cell wall composition has evolved and changed. Here stomatal cell wall composition was investigated in different vascular plant groups in attempt to understand their possible effect on stomatal function. Methods A renewed look at stomatal cell walls was attempted utilizing digitalized polar microscopy, confocal microscopy, histology and a numerical finite-elements simulation. The six species of vascular plants chosen for this study cover a broad structural, ecophysiological and evolutionary spectrum: ferns (Asplenium nidus and Platycerium bifurcatum) and angiosperms (Arabidopsis thaliana and Commelina erecta) with kidney-shaped stomata, and grasses (angiosperms, family Poaceae) with dumbbell-shaped stomata (Sorghum bicolor and Triticum aestivum). Key Results Three distinct patterns of cellulose crystallinity in stomatal cell walls were observed: Type I (kidney-shaped stomata, ferns), Type II (kidney-shaped stomata, angiosperms) and Type III (dumbbell-shaped stomata, grasses). The different stomatal cell wall attributes investigated (cellulose crystallinity, pectins, lignin, phenolics) exhibited taxon-specific patterns, with reciprocal substitution of structural elements in the end-walls of kidney-shaped stomata. According to a numerical bio-mechanical model, the end walls of kidney-shaped stomata develop the highest stresses during opening. Conclusions The data presented demonstrate for the first time the existence of distinct spatial patterns of varying cellulose crystallinity in guard cell walls. It is also highly intriguing that in angiosperms crystalline cellulose appears to have replaced lignin that occurs in the stomatal end-walls of ferns serving a similar wall strengthening function. Such taxon-specific spatial patterns of cell wall components could imply different biomechanical functions, which in turn could be a consequence of differences in environmental selection along the course of plant evolution. PMID:28158449

Large-Eddy Simulations of Noise Generation in Supersonic Jets at Realistic Engine Temperatures

NASA Astrophysics Data System (ADS)

Liu, Junhui; Corrigan, Andrew; Kailasanath, K.; Taylor, Brian

2015-11-01

Large-eddy simulations (LES) have been carried out to investigate the noise generation in highly heated supersonic jets at temperatures similar to those observed in high-performance jet engine exhausts. It is found that the exhaust temperature of high-performance jet engines can range from 1000K at an intermediate power to above 2000K at a maximum afterburning power. In low-temperature jets, the effects of the variation of the specific heat ratio as well as the radial temperature profile near the nozzle exit are small and are ignored, but it is not clear whether those effects can be also ignored in highly heated jets. The impact of the variation of the specific heat ratio is assessed by comparing LES results using a variable specific heat ratio with those using a constant specific heat ratio. The impact on both the flow field and the noise distributions are investigated. Because the total temperature near the nozzle wall can be substantially lower than the nozzle total temperature either due to the heating loss through the nozzle wall or due to the cooling applied near the wall, this lower wall temperature may impact the temperature in the shear layer, and thus impact the noise generation. The impact of the radial temperature profile on the jet noise generation is investigated by comparing results of lower nozzle wall temperatures with those of the adiabatic wall condition.

The genetic interaction network of CCW12, a Saccharomyces cerevisiae gene required for cell wall integrity during budding and formation of mating projections

PubMed Central

2011-01-01

Background Mannoproteins construct the outer cover of the fungal cell wall. The covalently linked cell wall protein Ccw12p is an abundant mannoprotein. It is considered as crucial structural cell wall component since in baker's yeast the lack of CCW12 results in severe cell wall damage and reduced mating efficiency. Results In order to explore the function of CCW12, we performed a Synthetic Genetic Analysis (SGA) and identified genes that are essential in the absence of CCW12. The resulting interaction network identified 21 genes involved in cell wall integrity, chitin synthesis, cell polarity, vesicular transport and endocytosis. Among those are PFD1, WHI3, SRN2, PAC10, FEN1 and YDR417C, which have not been related to cell wall integrity before. We correlated our results with genetic interaction networks of genes involved in glucan and chitin synthesis. A core of genes essential to maintain cell integrity in response to cell wall stress was identified. In addition, we performed a large-scale transcriptional analysis and compared the transcriptional changes observed in mutant ccw12Δ with transcriptomes from studies investigating responses to constitutive or acute cell wall damage. We identified a set of genes that are highly induced in the majority of the mutants/conditions and are directly related to the cell wall integrity pathway and cell wall compensatory responses. Among those are BCK1, CHS3, EDE1, PFD1, SLT2 and SLA1 that were also identified in the SGA. In contrast, a specific feature of mutant ccw12Δ is the transcriptional repression of genes involved in mating. Physiological experiments substantiate this finding. Further, we demonstrate that Ccw12p is present at the cell periphery and highly concentrated at the presumptive budding site, around the bud, at the septum and at the tip of the mating projection. Conclusions The combination of high throughput screenings, phenotypic analyses and localization studies provides new insight into the function of Ccw12p. A compensatory response, culminating in cell wall remodelling and transport/recycling pathways is required to buffer the loss of CCW12. Moreover, the enrichment of Ccw12p in bud, septum and mating projection is consistent with a role of Ccw12p in preserving cell wall integrity at sites of active growth. The microarray data produced in this analysis have been submitted to NCBI GEO database and GSE22649 record was assigned. PMID:21320323