Overexpression of vesicle-associated membrane protein PttVAP27-17 as a tool to improve biomass production and the overall saccharification yields in Populus trees

Abstract Background Bioconversion of wood into bioproducts and biofuels is hindered by the recalcitrance of woody raw material to bioprocesses such as enzymatic saccharification. Targeted modification of the chemical composition of the feedstock can improve saccharification but this gain is often abrogated by concomitant reduction in tree growth. Results In this study, we report on transgenic hybrid aspen (Populus tremula × tremuloides) lines that showed potential to increase biomass production both in the greenhouse and after 5 years of growth in the field. The transgenic lines carried an overexpression construct for Populus tremula × tremuloides vesicle-associated membrane protein (VAMP)-associated protein PttVAP27-17 that was selected from a gene-mining program for novel regulators of wood formation. Analytical-scale enzymatic saccharification without any pretreatment revealed for all greenhouse-grown transgenic lines, compared to the wild type, a 20–44% increase in the glucose yield per dry weight after enzymatic saccharification, even though it was statistically significant only for one line. The glucose yield after enzymatic saccharification with a prior hydrothermal pretreatment step with sulfuric acid was not increased in the greenhouse-grown transgenic trees on a dry-weight basis, but increased by 26–50% when calculated on a whole biomass basis in comparison to the wild-type control. Tendencies to increased glucose yields by up to 24% were present on a whole tree biomass basis after acidic pretreatment and enzymatic saccharification also in the transgenic trees grown for 5 years on the field when compared to the wild-type control. Conclusions The results demonstrate the usefulness of gene-mining programs to identify novel genes with the potential to improve biofuel production in tree biotechnology programs. Furthermore, multi-omic analyses, including transcriptomic, proteomic and metabolomic analyses, performed here provide a toolbox for future studies on the function of VAP27 proteins in plants.

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Overexpression of Mannitol-1-Phosphate Dehydrogenase Increases Mannitol Accumulation and Adds Protection Against Chilling Injury in Petunia

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.130.4.605 ◽

2005 ◽

Vol 130 (4) ◽

pp. 605-610 ◽

Cited By ~ 22

Author(s):

Yu-Jen Chiang ◽

C. Stushnoff ◽

A.E. McSay ◽

M.L. Jones ◽

H.J. Bohnert

Keyword(s):

Chilling Stress ◽

Chilling Injury ◽

Dry Weight ◽

Wild Type ◽

Gene Encoding ◽

E Coli ◽

Horticultural Crops ◽

Transgenic Lines ◽

And Control

Petunia ×hybrida (Hook) Vilm. cv. Mitchell was transformed with an E. coli gene encoding mannitol-1-phosphate dehydrogenase (mtlD). Four plant lines that grew on kanamycin and contained the mtlD transgene were identified. Two of these lines contained high levels of mannitol [high-mannitol lines M3 and M8; mean mannitol = 3.39 μmol·g-1 dry weight (DW)] compared to nontransformed wild-type plants (0.86 μmol·g-1 DW), while two lines had mannitol levels similar to wild-type plants (low-mannitol lines M2 and M9; mean mannitol = 1.05 μmol·g-1 DW). Transgenic and control plants were subjected to chilling stress (3 ± 0.5 °C day/0 ± 0.5 °C night, 12-hour photoperiod and 75% relative humidity) to evaluate the role of mannitol in chilling tolerance. Based upon foliage symptoms and membrane leakage after a 3-week chilling treatment, the high-mannitol containing lines, M3 and M8, were more tolerant of chilling stress than the low-mannitol containing transgenic lines, M2 and M9, and wild-type. Under nonchilling conditions mannitol was the only carbohydrate that differed among transgenic lines, but all carbohydrates were present. When subjected to chilling stress, mannitol levels dropped by 75%, sucrose by 52%, and inositol by 54% in the low-mannitol lines (M2 and M9). In M3 and M8, the high-mannitol lines, mannitol levels decreased by 36%, sucrose by 25%, and inositol by 56%, respectively. Raffinose increased 2- to 3-fold in all lines following exposure to low-temperature chilling stress. In the higher mannitol lines only 0.04% to 0.06% of the total osmotic potential generated from all solutes could be attributed to mannitol, thus its action is more like that of an osmoprotectant rather than an osmoregulator. This study demonstrates that metabolic engineering of osmoprotectant synthesis pathways can be used to improve stress tolerance in horticultural crops.

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Biomass production, transpiration rate and endogenous abscisic acid levels in grafts of flacca and wild-type tomato (Lycopersicon esculentum)

Functional Plant Biology ◽

10.1071/pp01263 ◽

2002 ◽

Vol 29 (11) ◽

pp. 1329 ◽

Cited By ~ 14

Author(s):

Guoxiong Chen ◽

S. Herman Lips ◽

Moshe Sagi

Keyword(s):

Plant Growth ◽

Biomass Production ◽

Transpiration Rate ◽

Loading Rate ◽

Dry Weight ◽

Wild Type ◽

Signalling Molecule ◽

Xylem Loading ◽

The Relationship ◽

Endogenous Aba

ABA is a plant signalling-molecule that plays a key role in regulating stomatal response, stress-tolerance responses, and coordinated growth of roots and shoots. Knowledge of the relationship between endogenous ABA level and plant growth is essential for improving plant growth and productivity. The wild-type tomato Rheinlands Ruhm (RR) and its flacca mutant were grafted in order to determine the relationship between endogenous ABA levels and biomass production. The flacca genotype is an ABA-deficient mutant, characterized by high stomatal conductance during the day or the night, high transpiration rate, marked tendency to wilt, and smaller size. Flacca scions grafted on to wild-type rootstock (Fs/Wr) exhibited higher ABA levels, lower transpiration rate, and higher water content than those of a control graft of flacca scion on flacca rootstock (Fs/Fr). Fs/Wr exhibited a lower ABA concentration, xylem exudate rate, ABA xylem-loading rate, and dry weight biomass in wild-type rootstock than in control grafts of wild-type scion on wild-type rootstock (Ws/Wr). Flacca rootstock grafted to wild-type scion (Ws/Fr) showed a higher ABA level, xylem exudation rate, ABA xylem-loading rate, dry weight biomass and length than grafts to flacca scion (Fs/Fr). Ws/Fr did not induce significant changes in wild-type scion as compared with Ws/Wr. In double grafts, flacca shoot fresh weight was significantly increased in flacca scion and wild-type scion grafted on to flacca rootstock (Fs + Ws/Fr) or wild-type rootstock (Fs + Ws/Wr). There was a significant linear relationship between biomass and ABA in scions (r=0.997, P=0.001). These results support the notion that ABA increases growth of tomato seedlings via improved stomatal control.

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A collection of genetically engineered Populus trees reveals wood biomass traits that predict glucose yield from enzymatic hydrolysis

10.1101/124396 ◽

2017 ◽

Author(s):

Sacha Escamez ◽

Madhavi Latha Gandla ◽

Marta Derba-Maceluch ◽

Sven-Olof Lundqvist ◽

Ewa J. Mellerowicz ◽

...

Keyword(s):

Enzymatic Hydrolysis ◽

Biomass Production ◽

Growth Parameters ◽

Genetically Engineered ◽

Wood Biomass ◽

Wild Type ◽

Glucose Yield ◽

Trade Offs ◽

Wild Type Clone ◽

Promising Source

AbstractWood represents a promising source of lignocellulosic biomass for the production of bio-based renewables, especially biofuels. However, woody feedstocks must be improved to become competitive against petroleum. We created a collection of Populus trees consisting of 40 genetically engineered lines to modify and to better understand wood biomass properties. A total of 65 traits were measured in these trees and in the corresponding wild-type clone, including growth parameters, wood anatomical and structural properties, cell wall composition and analytical saccharification. The relationships between saccharification of glucose and biomass traits were investigated using multivariate data analysis methods and mathematical modeling. To circumvent potential trade-offs between biomass production and saccharification potential, we also estimated the “total-wood glucose yield” (TWG) expected after pretreatment and 72h of enzymatic hydrolysis from whole trees. A mathematical model estimated TWG from a subset of 22 wood biomass traits with good predictivity (Q2 = 0.8), while saccharification of glucose could be predicted from seven biomass traits (Q2 = 0.49). Among the seven diagnostic traits for saccharification, four also affected biomass production, such as the ratio of S- to G-lignin which was beneficial for saccharification but detrimental for growth. The contents of various matrix polysaccharides appeared important for predicting both saccharification and TWG, including low abundance monosaccharides. In particular, fucose and mannose contents negatively correlated with TWG, apparently by negatively associating with biomass production. Both biomass production and saccharification, and hence TWG, negatively correlated with arabinose and rhamnose contents, suggesting that these low abundance monosaccharides represent markers/targets for improving feedstocks.

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Effects of an Autoregulatory Senescence-inhibitor Gene Construct on Nicotiana alata Link and Otto.

HortScience ◽

10.21273/hortsci.33.3.519d ◽

1998 ◽

Vol 33 (3) ◽

pp. 519d-519 ◽

Cited By ~ 4

Author(s):

Kenneth R. Schroeder ◽

Dennis P. Stimart

Keyword(s):

Agrobacterium Tumefaciens ◽

Plant Height ◽

Dry Weight ◽

Nicotiana Alata ◽

Wild Type ◽

Gene Encoding ◽

Delayed Senescence ◽

Gene Construct ◽

Shoot Dry Weight ◽

Inhibitor System

Nicotiana alata Link and Otto. was transformed via Agrobacterium tumefaciens encoding a senescence-specific promoter SAG12 cloned from Arabidopsis thaliana fused to a Agrobacterium tumefaciens gene encoding isopentenyl transferase (IPT) that catalyzes cytokinin synthesis. This was considered an autoregulatory senescence-inhibitor system. In 1996, we reported delayed senescence of intact flowers by 2 to 6 d and delayed leaf senescence of transgenic vs. wild-type N. alata. Further evaluations in 1997 revealed several other interesting effects of the SAG12-IPT gene construct. Measurement of chlorophyll content of mature leaves showed higher levels of both chlorophyll a and b in transgenic material under normal fertilization and truncated fertilization regimes. At 4 to 5 months of age transgenic plants expressed differences in plant height, branching, and dry weight. Plant height was reduced by 3 to 13 cm; branch counts increased 2 to 3 fold; and shoot dry weight increased up to 11 g over wild-type N. alata. These observations indicate the system is not tightly autoregulated and may prove useful to the floriculture industry for producing compact and more floriferous plants.

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Valorization of sugarcane bagasse by chemical pretreatment and enzyme mediated deconstruction

Scientific Reports ◽

10.1038/s41598-019-52347-7 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 7

Author(s):

Vihang S. Thite ◽

Anuradha S. Nerurkar

Keyword(s):

Cell Wall ◽

Sugarcane Bagasse ◽

Structural Changes ◽

Enzymatic Saccharification ◽

Dry Weight ◽

Gravimetric Analysis ◽

Chemical Pretreatment ◽

Cell Wall Degrading Enzymes ◽

First Time ◽

Soluble Components

Abstract After chemical pretreatment, improved amenability of agrowaste biomass for enzymatic saccharification needs an understanding of the effect exerted by pretreatments on biomass for enzymatic deconstruction. In present studies, NaOH, NH4OH and H2SO4 pretreatments effectively changed visible morphology imparting distinct fibrous appearance to sugarcane bagasse (SCB). Filtrate analysis after NaOH, NH4OH and H2SO4 pretreatments yielded release of soluble reducing sugars (SRS) in range of ~0.17–0.44%, ~0.38–0.75% and ~2.9–8.4% respectively. Gravimetric analysis of pretreated SCB (PSCB) biomass also revealed dry weight loss in range of ~25.8–44.8%, ~11.1–16.0% and ~28.3–38.0% by the three pretreatments in the same order. Release of soluble components other than SRS, majorly reported to be soluble lignins, were observed highest for NaOH followed by H2SO4 and NH4OH pretreatments. Decrease or absence of peaks attributed to lignin and loosened fibrous appearance of biomass during FTIR and SEM studies respectively further corroborated with our observations of lignin removal. Application of commercial cellulase increased raw SCB saccharification from 1.93% to 38.84%, 25.56% and 9.61% after NaOH, H2SO4 and NH4OH pretreatments. Structural changes brought by cell wall degrading enzymes were first time shown visually confirming the cell wall disintegration under brightfield, darkfield and fluorescence microscopy. The microscopic evidence and saccharification results proved that the chemical treatment valorized the SCB by making it amenable for enzymatic saccharification.

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Overexpression of Mucin 1 Suppresses the Therapeutical Efficacy of Disulfiram against Canine Mammary Tumor

Animals ◽

10.3390/ani11010037 ◽

2020 ◽

Vol 11 (1) ◽

pp. 37

Author(s):

Ying Zhao ◽

Zixiang Lin ◽

Zhaoyan Lin ◽

Chaoyu Zhou ◽

Gang Liu ◽

...

Keyword(s):

Mammary Tumor ◽

Mammary Tumors ◽

Expression Level ◽

Canine Mammary Tumor ◽

Wild Type ◽

Mucin 1 ◽

Mammary Tumor Cell ◽

Mammary Tumor Cell Line ◽

Canine Mammary Tumors ◽

Wild Type Control

Mucin 1 (MUC1), a transmembrane protein, is closely associated with the malignancy and metastasis of canine mammary tumors; however, the role of overexpressed MUC1 in the development of cancer cells and response to drug treatment remains unclear. To address this question, we developed a new canine mammary tumor cell line, CIPp-MUC1, with an elevated expression level of MUC1. In vitro studies showed that CIPp-MUC1 cells are superior in proliferation and migration than wild-type control, which was associated with the upregulation of PI3K, p-Akt, mTOR, Bcl-2. In addition, overexpression of MUC1 in CIPp-MUC1 cells inhibited the suppressing activity of disulfiram on the growth and metastasis of tumor cells, as well as inhibiting the pro-apoptotic effect of disulfiram. In vivo studies, on the other side, showed more rapid tumor growth and stronger resistance to disulfiram treatment in CIPp-MUC1 xenograft mice than in wild-type control. In conclusion, our study demonstrated the importance of MUC1 in affecting the therapeutical efficiency of disulfiram against canine mammary tumors, indicating that the expression level of MUC1 should be considered for clinical use of disulfiram or other drugs targeting PI3K/Akt pathway.

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Mating in Saccharomyces cerevisiae: The Role of the Pheromone Signal Transduction Pathway in the Chemotropic Response to Pheromone

Genetics ◽

10.1093/genetics/147.1.19 ◽

1997 ◽

Vol 147 (1) ◽

pp. 19-32 ◽

Cited By ~ 5

Author(s):

Kathrin Schrick ◽

Barbara Garvik ◽

Leland H Hartwell

Keyword(s):

Signal Transduction ◽

Map Kinase ◽

Transcriptional Activation ◽

Signal Transduction Pathway ◽

Transduction Pathway ◽

Wild Type ◽

Mating Partner ◽

Map Kinase Cascade ◽

Kinase Cascade ◽

Wild Type Control

Abstract The mating process in yeast has two distinct aspects. One is the induction and activation of proteins required for cell fusion in response to a pheromone signal; the other is chemotropism, i.e., detection of a pheromone gradient and construction of a fusion site available to the signaling cell. To determine whether components of the signal transduction pathway necessary for transcriptional activation also play a role in chemotropism, we examined strains with null mutations in components of the signal transduction pathway for diploid formation, prezygote formation and the chemotropic process of mating partner discrimination when transcription was induced downstream of the mutation. Cells mutant for components of the mitogen-activated protein (MAP) kinase cascade (ste5, ste20, ste11, ste7 or fus3 kss1) formed diploids at a frequency 1% that of the wild-type control, but formed prezygotes as efficiently as the wild-type control and showed good mating partner discrimination, suggesting that the MAP kinase cascade is not essential for chemotropism. In contrast, cells mutant for the receptor (ste2) or the β or γ subunit (ste4 and stel8) of the G protein were extremely defective in both diploid and prezygote formation and discriminated poorly between signaling and nonsignaling mating partners, implying that these components are important for chemotropism.

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A Cyanobacteria-Based Biofilm System for Advanced Brewery Wastewater Treatment

Applied Sciences ◽

10.3390/app11010174 ◽

2020 ◽

Vol 11 (1) ◽

pp. 174

Author(s):

Konstantinos P. Papadopoulos ◽

Christina N. Economou ◽

Athanasia G. Tekerlekopoulou ◽

Dimitris V. Vayenas

Keyword(s):

Wastewater Treatment ◽

Biomass Production ◽

Low Cost ◽

Oxygen Demand ◽

Dry Weight ◽

Surface Hydrophobicity ◽

Reactor Performance ◽

Brewery Wastewater ◽

Kjeldahl Nitrogen ◽

Supporting Materials

Algal/cyanobacterial biofilm photobioreactors provide an alternative technology to conventional photosynthetic systems for wastewater treatment based on high biomass production and easy biomass harvesting at low cost. This study introduces a novel cyanobacteria-based biofilm photobioreactor and assesses its performance in post-treatment of brewery wastewater and biomass production. Two different supporting materials (glass/polyurethane) were tested to investigate the effect of surface hydrophobicity on biomass attachment and overall reactor performance. The reactor exhibited high removal efficiency (over 65%) of the wastewater’s pollutants (chemical oxygen demand, nitrate, nitrite, ammonium, orthophosphate, and total Kjeldahl nitrogen), while biomass per reactor surface reached 13.1 and 12.8 g·m−2 corresponding to 406 and 392 mg·L−1 for glass and polyurethane, respectively, after 15 days of cultivation. The hydrophilic glass surface favored initial biomass adhesion, although eventually both materials yielded complete biomass attachment, highlighting that cell-to-cell interactions are the dominant adhesion mechanism in mature biofilms. It was also found that the biofilm accumulated up to 61% of its dry weight in carbohydrates at the end of cultivation, thus making the produced biomass a suitable feedstock for bioethanol production.

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Stress-induced expression of IPT gene in transgenic wheat reduces grain yield penalty under drought

Journal of Genetic Engineering and Biotechnology ◽

10.1186/s43141-021-00171-w ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Ailin Beznec ◽

Paula Faccio ◽

Daniel J. Miralles ◽

Leonor G. Abeledo ◽

Cecilia Decima Oneto ◽

...

Keyword(s):

Grain Yield ◽

Bread Wheat ◽

Transcriptional Control ◽

Field Conditions ◽

Wild Type ◽

Ipt Gene ◽

Receptor Like Kinase ◽

Biomass Components ◽

Yield Penalty ◽

Wild Type Control

Abstract Background The heterologous expression of isopentenyl transferase (IPT) under the transcriptional control of the senescence-associated receptor-like kinase (SARK) promoter delayed cellular senescence and, through it, increased drought tolerance in plants. To evaluate the effect of pSARK::IPT expression in bread wheat, six independent transgenic events were obtained through the biolistic method and evaluated transgene expression, phenology, grain yield and physiological biomass components in plants grown under both drought and well-irrigating conditions. Experiments were performed at different levels: (i) pots and (ii) microplots inside a biosafety greenhouse, as well as under (iii) field conditions. Results Two transgenic events, called TR1 and TR4, outperformed the wild-type control under drought conditions. Transgenic plants showed higher yield under both greenhouse and field conditions, which was positively correlated to grain number (given by more spikes and grains per spike) than wild type. Interestingly, this yield advantage of the transgenic events was observed under both drought and well-watered conditions. Conclusions The results obtained allow us to conclude that the SARK promoter-regulated expression of the IPT gene in bread wheat not only reduced the yield penalty produced by water stress but also led to improved productivity under well-watered conditions.

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