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

Abstract

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 increased 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.

Authors:
 [1];  [2];  [1];  [1];  [1];  [3];  [4];  [1];  [2];  [3]
  1. Univ. of Georgia, Athens, GA (United States)
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division
  3. Ohio Univ., Athens, OH (United States)
  4. Ohio Univ., Athens, OH (United States); Suzhou Inst. of Nano-Tech and Nano-Bionics (SINANO), Suzhou (China)
Publication Date:
Research Org.:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1260605
Alternate Identifier(s):
OSTI ID: 1265756
Grant/Contract Number:
PS02-06ER6430; MCB- 9874744; 526-HF02; AC05-00OR22725
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
PLoS ONE
Additional Journal Information:
Journal Volume: 9; Journal Issue: 12; Journal ID: ISSN 1932-6203
Publisher:
Public Library of Science
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; Cell walls; Plant cell walls; Cell fusion; Amino acid analysis; Cross-linking; Elastin; Cellulose

Citation Formats

Tan, Li, Pu, Yunqiao, Pattathil, Sivakumar, Avci, Utku, Qian, Jin, Arter, Allison, Chen, Liwei, Hahn, Michael G., Ragauskas, Arthur J., and Kieliszewski, Marcia J.. Changes in Cell Wall Properties Coincide with Overexpression of Extensin Fusion Proteins in Suspension Cultured Tobacco Cells. United States: N. p., 2014. Web. doi:10.1371/journal.pone.0115906.
Tan, Li, Pu, Yunqiao, Pattathil, Sivakumar, Avci, Utku, Qian, Jin, Arter, Allison, Chen, Liwei, Hahn, Michael G., Ragauskas, Arthur J., & Kieliszewski, Marcia J.. Changes in Cell Wall Properties Coincide with Overexpression of Extensin Fusion Proteins in Suspension Cultured Tobacco Cells. United States. doi:10.1371/journal.pone.0115906.
Tan, Li, Pu, Yunqiao, Pattathil, Sivakumar, Avci, Utku, Qian, Jin, Arter, Allison, Chen, Liwei, Hahn, Michael G., Ragauskas, Arthur J., and Kieliszewski, Marcia J.. Tue . "Changes in Cell Wall Properties Coincide with Overexpression of Extensin Fusion Proteins in Suspension Cultured Tobacco Cells". United States. doi:10.1371/journal.pone.0115906. https://www.osti.gov/servlets/purl/1260605.
@article{osti_1260605,
title = {Changes in Cell Wall Properties Coincide with Overexpression of Extensin Fusion Proteins in Suspension Cultured Tobacco Cells},
author = {Tan, Li and Pu, Yunqiao and Pattathil, Sivakumar and Avci, Utku and Qian, Jin and Arter, Allison and Chen, Liwei and Hahn, Michael G. and Ragauskas, Arthur J. and Kieliszewski, Marcia J.},
abstractNote = {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 increased 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.},
doi = {10.1371/journal.pone.0115906},
journal = {PLoS ONE},
number = 12,
volume = 9,
place = {United States},
year = {Tue Dec 23 00:00:00 EST 2014},
month = {Tue Dec 23 00:00:00 EST 2014}
}

Journal Article:
Free Publicly Available Full Text
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Cited by: 4works
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  • Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp 4 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 andmore » increased 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. These data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.« less
  • The pathway of hydroxyproline-containing proteins to the cell wall and to the growth medium in suspension-cultured Acer pseudoplatanus cells is traced by following the kinetics of the transfer of protein-bound /sup 14/C-hydroxyproline into various fractions, and by comparing the hydroxyproline-arabinoside profiles of these fractions after alkaline hydrolysis. Hydroxyproline-rich protein passes directly from a membrane-bound compartment in the cytoplasm to the cell wall, not via an intermediate salt-soluble pool in the wall. There are at least three hydroxyproline-containing glycoproteins in the cell wall. One which possesses mono-, tri-, and tetraarabinoside side chains accounts for over 90% of the total hydroxyproline. Thismore » glycoprotein is ''extensin.'' The hydroxyproline-containing proteins secreted into the medium have a glycosylation pattern markedly different from that of the major cell wall glycoprotein. It appears that there is little or no wall-like extensin in the medium. Approximately half of the protein-bound hydroxyproline secreted into the medium is linked to an arabinogalactan. This linkage is also found in a particulate wall protein precursor fraction from the cytoplasm, but only trace amounts can be detected in the cell wall.« less
  • The partial purification and characterization of cell wall polysaccharides isolated from suspension-cultured Douglas fir (Pseudotsuga menziesii) cells are described. Extraction of isolated cell walls from 1.0 M LiCl solubilized pectic polysaccharides with glycosyl-linkage compositions similar to those of rhamnogalacturonans I and II, pectic polysaccharides isolated from walls of suspension-cultured sycamore cells. Treatment of LiCl-extracted Douglas fir walls with an endo-..cap alpha..-1,4-polygalacturonase released only small, additional amounts of pectic polysaccharide, which had a glycosyl-linkage composition similar to that of rhamnogalacturonan I. Xyloglucan oligosaccharides were released from the endo-..cap alpha..-1,4-polygalacturonase-treated walls by treatment with an endo-..beta..-1,4-glucanase. These oligosaccharides included hepta- and nonasaccharidesmore » similar or identical to those released from sycamore cell walls by the same enzyme, and structurally related octa- and decasaccharides similar to those isolated from various angiosperms. Finally, additional xyloglucan and small amounts of xylan were extracted from the endo-..beta..-1,4-glucanase-treated walls by 0.5 N NaOH. The xylan resembled that extracted by NaOH from dicot cell walls in that it contained 2,4- but not 3,4-linked xylosyl residues. In this study, a total of 15% of the cell wall was isolated as pectic material, 10% as xyloglucan, and less than 1% as xylan. The noncellulosic polysaccharides accounted for 25% of the cell walls, cellulose for 23%, protein for 34%, and ash for 5%, for a total of 88% of the cell wall.« less
  • Inhibition of cell growth and accumulation of Cd-binding peptide were measured in cultured tobacco cells exposed to buthionine sulfoximine. This inhibitor of glutathione metabolism caused little or no reduction of growth (at 0.1 millimolar) in the absence of Cd, but growth was greatly reduced in cultures exposed to buthionine sulfoximine and greater than or equal to 22 micromolar Cd. Decreased cell growth was directly correlated with decreased levels of Cd-binding peptide and increased level of what is thought to be free Cd. Zinc inhibited growth of tobacco cells only at the highest levels examined (900-1800 micromolar Zn), but buthionine sulfoximinemore » had no additional significantly effect. Similar results were observed for Cu (45-90 micromolar). Results suggest that synthesis of plant Cd-peptide involves ..gamma..-glutamylcysteine synthetase or a related enzyme and that Zn accumulation in tobacco cells does not cause formation of significant Cd-peptide ligand.« less
  • Cultured tobacco cells (Nicotiana tabacum L. cv Wisconsin 38) adapted to grow in medium containing high levels of NaCl or polyethylene glycol (PEG) produce several new or enhanced polypeptide bands on sodium dodecyl sulfate-polyarylamide gel electrophoresis. The intensities of some of the polypeptide bands increase with increasing levels of NaCl adaptation, while the intensities of other polypeptide bands are reduced. Synthesis of 26-kilodalton polypeptide(s) occurs at two different periods during culture growth of NaCl adapted cells. Unadapted cells also incorporate /sup 35/S into a 26-kilodalton polypeptide during the later stage of culture growth beginning at midlog phase. The 26-kilodalton polypeptidesmore » from adapted and unadapted cells have similar partial proteolysis peptide maps and are immunologically cross-reactive. During adaptation to NaCl, unadapted cells synthesize and accumulate a major 26-kilodalton polypeptide, and the beginning of synthesis corresponds to the period of osmotic adjustment and culture growth. From their results, the authors suggest an involvement of the 26-kilodalton polypeptide in the adaptation of cultured tobacco cells to NaCl and water stress. 38 references, 11 figures, 2 tables.« less