| 1. |
- Endo, Satoshi, et al.
(författare)
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Instability of C154Y variant of aldo-keto reductase 1C3
- 2017
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Ingår i: Chemico-Biological Interactions. - : Elsevier. - 0009-2797 .- 1872-7786. ; 276, s. 194-202
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Tidskriftsartikel (refereegranskat)abstract
- Aldo-keto reductase (AKR) 1C3 is a cytosolic enzyme that metabolizes steroids, prostaglandins, toxic aldehydes and drugs. Recently, some nonsynonymous single nucleotide polymorphisms of AKR1C3 have been suggested to impact steroid and drug metabolism. In this study, we examined the effects of C154Y and L159V variants of AKR1C3 on stability and function of the enzyme. Both variants had been detected in patients with the neurodegenerative disease amyotrophic lateral sclerosis. Recombinant wild-type (WT), C154Y and L159V enzymes were similar in specific activity, but C154Y displayed much lower thermostability than WT and L159V. C154Y was inactivated by 10-min incubation at >25 °C, and about 90% of its activity was lost at 40 °C. Differential scanning fluorimetry revealed that Tm (thermal denaturation midpoint) of C154Y was lower than that of WT. In order to study the cause of thermosensitivity of C154Y, we prepared C154F and C154S mutant AKR1C3s. Like C154Y, C154F was highly sensitive to thermal inactivation, whereas C154S showed almost the same thermostability as WT. The C154F and C154Y variants induced secondary and tertiary structural changes in AKR1C3 at 40 °C as reflected by their altered circular dichroism and 8-anilinonaphthalene-1-sulfonate fluorescence characteristics. These results suggest that the replacement of C154 with a residue possessing a bulky aromatic side-chain impairs the folding of the α-helix containing C154 and its neighboring secondary structures, leading to low thermostability of AKR1C3. AKR1C3 metabolizes cytotoxic 4-oxo-2-nonenal into a less toxic metabolite, and overexpression of WT in HEK293 cells alleviated the 4-oxo-2-nonenal-induced cytotoxicity. In contrast, the overexpression of C154Y in the cells did not show such a significant protective effect, suggesting that C154Y is unstable in cells.
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| 2. |
- Endo, Satoshi, et al.
(författare)
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Transient transformation and RNA silencing in Zinnia tracheary element differentiating cell cultures.
- 2008
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Ingår i: The Plant journal : for cell and molecular biology. - 1365-313X. ; 53:5, s. 864-75
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Tidskriftsartikel (refereegranskat)abstract
- The Zinnia elegans cell culture system is a robust and physiologically relevant in vitro system for the study of xylem formation. Freshly isolated mesophyll cells of Zinnia can be hormonally induced to semisynchronously transdifferentiate into tracheary elements (TEs). Although the system has proven to be valuable, its utility is diminished by the lack of an efficient transformation protocol. We herein present a novel method to introduce DNA/RNA efficiently into Zinnia cells by electroporation-based transient transformation. Using reporter gene plasmids, we optimized the system for efficiency of transformation and ability for the transformed cells to transdifferentiate into TEs. Optimal conditions included a partial digestion of the cell walls by pectolyase, a low voltage and high capacitance electrical pulse and an optimal medium to maintain cell viability during transformation. Beyond the simple expression of a reporter protein in Zinnia cells, we extended our protocol to subcellular protein targeting, simultaneous co-expression of several reporter proteins and promoter-activity monitoring during TE differentiation. Most importantly, we tested the system for double-stranded RNA (dsRNA)-induced RNA silencing. By introducing in vitro-synthesized dsRNAs, we were able to phenocopy the Arabidopsis cellulose synthase (CesA) mutants that had defects in secondary cell-wall synthesis. Suppressing the expression ofZinnia CesA homologues resulted in an increase of abnormal TEs with aberrant secondary walls. Our electroporation-based transient transformation protocol provides the suite of tools long required for functional analysis and developmental studies at single cell levels.
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| 3. |
- Hale, Sarah E., et al.
(författare)
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Sorption of the monoterpenes alpha-pinene and limonene to carbonaceous geosorbents including biochar
- 2015
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Ingår i: Chemosphere. - : Elsevier BV. - 0045-6535 .- 1879-1298. ; 119, s. 881-888
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Tidskriftsartikel (refereegranskat)abstract
- The sorption of two monoterpenes, alpha pinene and limonene to the carbonaceous geosorbents graphite, bituminous coal, lignite coke, biochar and Pahokee peat was quantified. Polyethylene (PE) passive samplers were calibrated for the first time for these compounds by determining the PE-water partitioning coefficients and used as a tool to determine sorption to the carbonaceous geosorbents. Log KPE-water values were 3.49 +/- 0.58 for alpha pinene and 4.08 +/- 0.27 for limonene. The sorption of limonene to all materials was stronger than that for a pinene (differences of 0.2-13 log units between distribution coefficients for the monoterpenes). Placing K-d values in increasing order for a pinene gave biochar approximate to Pahokee peat approximate to bituminous coal approximate to lignite coke < graphite. For limonene the order was: Pahokee peat approximate to biochar approximate to bituminous coal < graphite approximate to lignite coke. Micropore (defined as pores <1.5 nm) and nanopore surface area (defined as pores 1.5 nm to 50 nm) normalised carbonaceous geosorbent-water distribution coefficients were also calculated. There was no clear correlation of these distribution coefficients with SA. Elemental composition was used to assess the degree of condensation (or alteration) of the carbonaceous geosorbents. The degree of carbonisation increased in the order; Pahokee peat < lignite coke < bituminous coal < biochar < graphite, however this was not correlated with an increase in the experimental distribution coefficients.
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| 4. |
- Kasaba, Yasumasa, et al.
(författare)
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Mission Data Processor Aboard the BepiColombo Mio Spacecraft : Design and Scientific Operation Concept
- 2020
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Ingår i: Space Science Reviews. - : Springer Science and Business Media LLC. - 0038-6308 .- 1572-9672. ; 216:3
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Forskningsöversikt (refereegranskat)abstract
- BepiColombo Mio, also known as the Mercury Magnetospheric Orbiter (MMO), is intended to conduct the first detailed study of the magnetic field and environment of the innermost planet, Mercury, alongside the Mercury Planetary Orbiter (MPO). This orbiter has five payload groups; the MaGnetic Field Investigation (MGF), the Mercury Plasma Particle Experiment (MPPE), the Plasma Wave Investigation (PWI), the Mercury Sodium Atmosphere Spectral Imager (MSASI), and the Mercury Dust Monitor (MDM). These payloads operate through the Mission Data Processor (MDP) that acts as an integrated system for Hermean environmental studies by the in situ observation of charged and energetic neutral particles, magnetic and electric fields, plasma waves, dust, and the remote sensing of radio waves and exospheric emissions. The MDP produces three kinds of coordinated data sets: Survey (L) mode for continuous monitoring, Nominal (M) mode for standard analyses of several hours in length (or more), and Burst (H) mode for analysis based on 4-20-min-interval datasets with the highest cadence. To utilize the limited telemetry bandwidth, nominal- and burst-mode data sets are partially downlinked after selections of data based on L- or L/M-mode data, respectively. Burst-mode data can be taken at preset timings, or by onboard automatic triggering. The MDP functions are implemented and tested on the ground as well as cruising spacecraft; they are responsible for conducting full scientific operations aboard spacecraft.
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| 5. |
- Kushwah, Sunita, et al.
(författare)
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Arabidopsis XTH4 and XTH9 Contribute to Wood Cell Expansion and Secondary Wall Formation(1)([OPEN])
- 2020
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Ingår i: Plant Physiology. - : American Society of Plant Science. - 0032-0889 .- 1532-2548. ; 182:4, s. 1946-1965
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Tidskriftsartikel (refereegranskat)abstract
- Xyloglucan is the major hemicellulose of dicotyledon primary cell walls, affecting the load-bearing framework with the participation of xyloglucan endo-transglycosylase/hydrolases (XTHs). We used loss- and gain-of function approaches to study functions of XTH4 and XTH9 abundantly expressed in cambial regions during secondary growth of Arabidopsis (Arabidopsis thaliana). In secondarily thickened hypocotyls, these enzymes had positive effects on vessel element expansion and fiber intrusive growth. They also stimulated secondary wall thickening but reduced secondary xylem production. Cell wall analyses of inflorescence stems revealed changes in lignin, cellulose, and matrix sugar composition indicating an overall increase in secondary versus primary walls in mutants, indicative of higher xylem production compared with the wild type (since secondary walls were thinner). Intriguingly, the number of secondary cell wall layers compared with the wild type was increased in xth9 and reduced in xth4, whereas the double mutant xth4x9 displayed an intermediate number of layers. These changes correlated with specific Raman signals from the walls, indicating changes in lignin and cellulose. Secondary walls were affected also in the interfascicular fibers, where neither XTH4 nor XTH9 was expressed, indicating that these effects were indirect. Transcripts involved in secondary wall biosynthesis and cell wall integrity sensing, including THESEUS1 and WALL ASSOCIATED KINASE2, were highly induced in the mutants, indicating that deficiency in XTH4 and XTH9 triggers cell wall integrity signaling, which, we propose, stimulates xylem cell production and modulates secondary wall thickening. Prominent effects of XTH4 and XTH9 on secondary xylem support the hypothesis that altered xyloglucan affects wood properties both directly and via cell wall integrity sensing.
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| 6. |
- Okada, Tatsuaki, et al.
(författare)
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Thermal Infrared Imaging Experiments of C-Type Asteroid 162173 Ryugu on Hayabusa2
- 2017
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Ingår i: Space Science Reviews. - : Springer Science and Business Media LLC. - 0038-6308 .- 1572-9672. ; 208:1-4, s. 255-286
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Tidskriftsartikel (refereegranskat)abstract
- The thermal infrared imager TIR onboard Hayabusa2 has been developed to investigate thermo-physical properties of C-type, near-Earth asteroid 162173 Ryugu. TIR is one of the remote science instruments on Hayabusa2 designed to understand the nature of a volatile-rich solar system small body, but it also has significant mission objectives to provide information on surface physical properties and conditions for sampling site selection as well as the assessment of safe landing operations. TIR is based on a two-dimensional uncooled micro-bolometer array inherited from the Longwave Infrared Camera LIR on Akatsuki (Fukuhara et al., 2011). TIR takes images of thermal infrared emission in 8 to 12 μm with a field of view of 16×12∘16×12∘ and a spatial resolution of 0.05∘0.05∘ per pixel. TIR covers the temperature range from 150 to 460 K, including the well calibrated range from 230 to 420 K. Temperature accuracy is within 2 K or better for summed images, and the relative accuracy or noise equivalent temperature difference (NETD) at each of pixels is 0.4 K or lower for the well-calibrated temperature range. TIR takes a couple of images with shutter open and closed, the corresponding dark frame, and provides a true thermal image by dark frame subtraction. Data processing involves summation of multiple images, image processing including the StarPixel compression (Hihara et al., 2014), and transfer to the data recorder in the spacecraft digital electronics (DE). We report the scientific and mission objectives of TIR, the requirements and constraints for the instrument specifications, the designed instrumentation and the pre-flight and in-flight performances of TIR, as well as its observation plan during the Hayabusa2 mission.
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| 7. |
- Ratke, Christine, et al.
(författare)
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Populus GT43 family members group into distinct sets required for primary and secondary wall xylan biosynthesis and include useful promoters for wood modification
- 2015
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Ingår i: Plant Biotechnology Journal. - : Wiley. - 1467-7644 .- 1467-7652. ; 13:1, s. 26-37
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Tidskriftsartikel (refereegranskat)abstract
- The plant GT43 protein family includes xylosyltransferases that are known to be required for xylan backbone biosynthesis, but have incompletely understood specificities. RT-qPCR and histochemical (GUS) analyses of expression patterns of GT43 members in hybrid aspen, reported here, revealed that three clades of the family have markedly differing specificity towards secondary wall-forming cells (wood and extraxylary fibres). Intriguingly, GT43A and B genes (corresponding to the Arabidopsis IRX9 clade) showed higher specificity for secondary-walled cells than GT43C and D genes (IRX14 clade), although both IRX9 and IRX14 are required for xylosyltransferase activity. The remaining genes, GT43E, F and G (IRX9-L clade), showed broad expression patterns. Transient transactivation analyses of GT43A and B reporters demonstrated that they are activated by PtxtMYB021 and PNAC085 (master secondary wall switches), mediated in PtxtMYB021 activation by an AC element. The high observed secondary cell wall specificity of GT43B expression prompted tests of the efficiency of its promoter (pGT43B), relative to the CaMV 35S (35S) promoter, for overexpressing a xylan acetyl esterase (CE5) or downregulating REDUCED WALL ACETYLATION (RWA) family genes and thus engineering wood acetylation. CE5 expression was weaker when driven by pGT43B, but it reduced wood acetyl content substantially more efficiently than the 35S promoter. RNAi silencing of the RWA family, which was ineffective using 35S, was achieved when using GT43B promoter. These results show the utility of the GT43B promoter for genetically engineering properties of wood and fibres.
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| 8. |
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| 9. |
- Takahashi Schmidt, Junko, et al.
(författare)
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KORRIGAN1 and its Aspen Homolog PttCel9A1 Decrease Cellulose Crystallinity in Arabidopsis Stems
- 2009
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Ingår i: Plant and Cell Physiology. - : Oxford University Press (OUP). - 0032-0781 .- 1471-9053. ; 50:6, s. 1099-1115
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Tidskriftsartikel (refereegranskat)abstract
- KORRIGAN1 (KOR1) is a membrane-bound cellulase implicated in cellulose biosynthesis. PttCel9A1 from hybrid aspen (Populus tremula L. tremuloides Michx.) has high sequence similarity to KOR1 and we demonstrate here that it complements kor1-1 mutants, indicating that it is a KOR1 ortholog. We investigated the function of PttCel9A1/KOR1 in Arabidopsis secondary growth using transgenic lines expressing 35S::PttCel9A1 and the KOR1 mutant line irx2-2. The presence of elevated levels of PttCel9A1/KOR1 in secondary walls of 35S::PttCel9A1 lines was confirmed by in muro visualization of cellulase activity. Compared with the wild type, 35S::PttCel9A1 lines had higher trifluoroacetic acid (TFA)-hydrolyzable glucan contents, similar Updegraff cellulose contents and lower cellulose crystallinity indices, as determined by C-13 solid-state nuclear magnetic resonance (NMR) spectroscopy. irx2-2 mutants had wild-type TFA-hydrolyzable glucan contents, but reduced Updegraff cellulose contents and higher than wild-type cellulose crystallinity indices. The data support the hypothesis that PttCel9A1/KOR1 activity is present in cell walls, where it facilitates cellulose biosynthesis in a way that increases the amount of non-crystalline cellulose.
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