Search

Among the components of the blood-brain barrier (BBB), endothelial cells (ECs) play an important role in supplying limited materials, especially glucose, to the brain. However, the mechanism by which glucose is metabolized in brain ECs is still elusive. To address this topic, we assessed the metabolic signature of glucose utilization using live-cell metabolic assays and liquid chromatography-tandem mass spectrometry metabolomic analysis. We found that brain ECs are highly dependent on aerobic glycolysis, generating lactate as its final product with minimal consumption of glucose. Glucose treatment decreased the oxygen consumption rate in a dose-dependent manner, indicating the Crabtree effect. Moreover, when glycolysis was inhibited, brain ECs showed impaired permeability to molecules utilizing transcellular pathway. In addition, we found that the blockade of glycolysis in mouse brain with 2-deoxyglucose administration resulted in decreased transcellular permeability of the BBB. In conclusion, utilizing glycolysis in brain ECs has critical roles in the maintenance and permeability of the BBB. Overall, we could conclude that brain ECs are highly glycolytic, and their energy can be used to maintain the transcellular permeability of the BBB.

RNA-binding proteins (RBPs) play essential roles in diverse cellular processes through post-transcriptional regulation of RNAs. The subcellular localization of RBPs is thus under tight control, the breakdown of which is associated with aberrant cytoplasmic accumulation of nuclear RBPs such as TDP-43 and FUS, well-known pathological markers for amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD). Here, we report in Drosophila model for ALS/FTD that nuclear accumulation of a cytoplasmic RBP Staufen may be a new pathological feature. We found that in Drosophila C4da neurons expressing PR36, one of the arginine-rich dipeptide repeat proteins (DPRs), Staufen accumulated in the nucleus in Importin- and RNA-dependent manner. Notably, expressing Staufen with exogenous NLS—but not with mutated endogenous NLS—potentiated PR-induced dendritic defect, suggesting that nuclear-accumulated Staufen can enhance PR toxicity. PR36 expression increased Fibrillarin staining in the nucleolus, which was enhanced by heterozygous mutation of stau (stau+/−), a gene that codes Staufen. Furthermore, knockdown of fib, which codes Fibrillarin, exacerbated retinal degeneration mediated by PR toxicity, suggesting that increased amount of Fibrillarin by stau+/− is protective. stau+/− also reduced the amount of PR-induced nuclear-accumulated Staufen and mitigated retinal degeneration and rescued viability of flies expressing PR36. Taken together, our data show that nuclear accumulation of Staufen in neurons may be an important pathological feature contributing to the pathogenesis of ALS/FTD.

The hydrophobicity of film surfaces can be enhanced by introducing wrinkling and buckling, and the wrinkling method in this study is a facile way to induce various microstructures including wrinkles and buckles in film surfaces. When buckles formed, the water contact angle parallel to the wrinkled direction (θ//) changed dramatically with strain, but the contact angle perpendicular to the wrinkled direction did not change much. It was found that the buckle structural feature was the key characteristics affecting the change in contact angle, θ//. A dimensionless number, the ratio of buckle height to spacing (Hb/Sb) was introduced to determine the relationship between the structural feature and the contact angle, θ//. The contact angle, θ// strongly depended on Hb/Sb. The thickest film (hf = 605 nm) at the highest strain (ɛ = 30%) and the highest Hb/Sb formed many large buckles, and exhibited super-hydrophobicity (θ// = 169 ± 10°).

MDC1, a mediator of DNA damage response, recruits other repair proteins on double-strand break (DSB) sites. MDC1 is necessary for activating checkpoint kinases Chk1 and Chk2. It is unclear whether Chk1 interacts with MDC1. MDC1 also comprises many discrete domains. The role of the proline-serine-threonine (PST)-repeat domain of MDC1 in the DNA damage response is unclear. Here, we showed that MDC1 directly binds Chk1 through this PST-repeat region. Phosphorylation of Chk1 by ionizing radiation (IR) also required this PST-repeat domain. Degradation of intact MDC1 was accelerated depending on the PST-repeat domain after IR exposure. In the IR damage response, the PST-repeat-deleted MDC1 levels remained elevated with slow degradation. This abnormal regulation of MDC1 was F-box- and WD40 repeat-containing 7 (FBXW7)-dependent. The mutation of lysine 1413 within the PST-repeat of MDC1 deregulated MDC1 with or without damage. K1413R mutant and PST-deleted MDC1 displayed reduced ability to repair the damaged genome post-IR exposure. These results provide that the PST domain of MDC1 is involved in Chk1 and DNA repair activation. The findings suggest new insights into how MDC1 connects the checkpoint and DNA repair in the DNA damage response.

Acrylic pressure sensitive adhesives (PSAs) were prepared by UV polymerization under varying curing conditions of both fast and slow curing, employing high- and low-intensity UV radiation, respectively. The influences of curing conditions and isobornyl acrylate (IBOA) content on PSA performance were comprehensively investigated by measurement of their rheological, thermal, and adhesive properties. In particular, rheological characterization was accomplished by several analytical methods, such as in situ UV rheology, frequency sweep, stress relaxation, and temperature ramp tests, to understand the effect of the UV curing process and IBOA content on the viscoelastic behavior of acrylic PSAs. The slow-cured samples were observed to form more tightly crosslinked networks compared to the fast-cured. On the other hand, at high loading levels of IBOA, in the case of slow curing, the sample exhibited a contrasting trend, having the shortest stress relaxation time and the highest energy dissipation; this was due to molecular chain scission occurring in the crosslinked polymer during UV polymerization. Consequently, we successfully demonstrated the influence of monomer composition of acrylic PSAs, and that of curing conditions employed in UV polymerization. This study provides valuable insights for the development of crosslinked polymer networks of acrylic PSAs for flexible display applications.

The embryo phenotyping of genetic murine model is invaluable when investigating functions of genes underlying embryonic development and birth defect. Although traditional imaging technologies such as ultrasound are very useful for evaluating phenotype of murine embryos, the use of advanced techniques for phenotyping is desirable to obtain more information from genetic research. This letter tests the feasibility of optical coherence tomography (OCT) as a high-throughput phenotyping tool for murine embryos. Three-dimensional OCT imaging is performed for live and cleared mouse embryos in the late developmental stage (embryonic day 17.5). By using a dynamic focusing method and OCT angiography (OCTA) approach, our OCT imaging of the embryo exhibits rapid and clean visualization of organ structures deeper than 5 mm and complex microvasculature of perfused blood vessels in the murine embryonic body. This demonstration suggests that OCT imaging can be useful for comprehensively assessing embryo anatomy and angiography of genetically engineered mice.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes the coronavirus disease (COVID-19), is currently infecting millions of people worldwide and is causing drastic changes in people’s lives. Recent studies have shown that neurological symptoms are a major issue for people infected with SARS-CoV-2. However, the mechanism through which the pathological effects emerge is still unclear. Brain endothelial cells (ECs), one of the components of the blood–brain barrier, are a major hurdle for the entry of pathogenic or infectious agents into the brain. They strongly express angiotensin converting enzyme 2 (ACE2) for its normal physiological function, which is also well-known to be an opportunistic receptor for SARS-CoV-2 spike protein, facilitating their entry into host cells. First, we identified rapid internalization of the receptor-binding domain (RBD) S1 domain (S1) and active trimer (Trimer) of SARS-CoV-2 spike protein through ACE2 in brain ECs. Moreover, internalized S1 increased Rab5, an early endosomal marker while Trimer decreased Rab5 in the brain ECs. Similarly, the permeability of transferrin and dextran was increased in S1 treatment but decreased in Trimer, respectively. Furthermore, S1 and Trimer both induced mitochondrial damage including functional deficits in mitochondrial respiration. Overall, this study shows that SARS-CoV-2 itself has toxic effects on the brain ECs including defective molecular delivery and metabolic function, suggesting a potential pathological mechanism to induce neurological signs in the brain.

The responses of central nervous system (CNS) cells such as neurons and glia in neurodegenerative diseases (NDs) suggest that regulation of neuronal and glial functions could be a strategy for ND prevention and/or treatment. However, attempts to develop such therapeutics for NDs have been hindered by the challenge of blood-brain barrier (BBB) permeability and continued constitutive neuronal loss. These limitations indicate the need for additional perspectives for the prevention/treatment of NDs. In particular, the disruption of the blood-brain barrier (BBB) that accompanies NDs allows brain infiltration by peripheral factors, which may stimulate innate immune responses involved in the progression of neurodegeneration. The accumulation of blood factors like thrombin, fibrinogen, c-reactive protein (CRP) and complement components in the brain has been observed in NDs and may activate the innate immune system in the CNS. Thus, strengthening the integrity of the BBB may enhance its protective role to attenuate ND progression and functional loss. In this review, we describe the innate immune system in the CNS and the contribution of blood factors to the role of the CNS immune system in neurodegeneration and neuroprotection.

We conducted a preliminary study on fiber structural development in the high-speed melt spinning of environmentally friendly polyethylene terephthalate (Ti-PET) synthesized with 25 ppm of titanium-based catalyst, which was compared with conventional PET (Sb-PET) synthesized with 260 ppm of antimony-based catalyst. Gel permeation chromatography of Ti- and Sb-PET resins of intrinsic viscosity 0.63 confirmed that both resins have similar molecular weights and distributions. However, differential scanning calorimetry revealed that the Ti-PET resin exhibited a lower melt&ndash, crystallization peak and isothermal melt-crystallization rate than the Sb-PET resin. High-speed melt spinning of the Ti- and Sb-PET was possible up to a spinning velocity of 6 km/min. Two-dimensional wide-angle X-ray diffraction analyses showed that the molecular orientation of the obtained as-spun Ti- and Sb-PET fibers increased with spinning velocity, and a highly oriented, crystalline structure by orientation-induced crystallization started to appear from 5 km/min. Notably, Ti-PET fibers showed a lower degree of crystalline structural development and lower tensile strength compared with Sb-PET fibers under the high-speed spinning conditions. Our results suggest that the catalyst in PET resins can act as nucleating agents in thermal- and orientation-induced crystallization, and that differences in catalyst content can influence PET fiber structure development under extreme conditions in high-speed melt spinning.

Accumulation of RNA in the nucleus is one of the pathological features of C9orf72-associated amyotrophic lateral sclerosis and frontotemporal dementia (C9-ALS/FTD), yet its potential toxic cellular consequences remain largely undefined. RNA accumulated in the nucleus may interact with and increase nuclear localization of RNA-binding proteins (RBPs). Here, we show in C9-ALS/FTD Drosophila melanogaster model that Staufen, a double-stranded RBP normally localized in cytoplasm, accumulates in the nucleus, which is in contrast to many nuclear-localized RBPs, such as TDP-43 and FUS, whose cytoplasmic accumulation is thought to be a pathological hallmark of ALS/FTD. We found that in Drosophila neurons expressing arginine-rich dipeptide repeat proteins (DPRs), Staufen accumulated in the nucleus in an RNA-dependent manner. In the nucleus, Staufen localized closely to, and potentially interacts with, heterochromatin and nucleolus in Drosophila C4 da neurons expressing poly(PR), a proline-arginine (PR) DPR. PR toxicity in C4 da neurons increased Fibrillarin staining in the nucleolus, which was enhanced by stau heterozygous mutation. Furthermore, knockdown of fib exacerbated retinal degeneration mediated by PR toxicity, which suggests that increased amount of Fibrillarin by stau heterozygous mutation is protective. Heterozygotic mutation of stau could also mitigate retinal degeneration and rescue viability of flies exhibiting PR toxicity. Taken together, our data show that nuclear accumulation of cytoplasmic protein, such as Staufen, may also be an important pathological feature of C9-ALS/FTD.Author summaryCytoplasmic accumulation of nuclear RNA-binding proteins (RBPs) is one of the common pathological features of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). In C9orf72-associated ALS/FTD fly model, we found that Staufen, a double-stranded (ds) RBP normally localized mostly in cytoplasm, accumulates in the nucleus in an RNA-dependent manner. Next, we checked wherein the nucleus Staufen accumulates and found that Staufen partially co-localizes with heterochromatin and nucleolus. Interestingly, the expression of Fibrillarin, a nucleolar protein, was significantly increased by C9orf72-derived PR toxicity and further augmented by reduction in stau dosage, the gene encoding Staufen. When we knocked down fib, the gene encoding Fibrillarin, PR-induced retinal degeneration was exacerbated. This indicates that increased Fibrillarin expression by stau dosage reduction is protective. Furthermore, when we reduced stau dosage in flies presenting PR toxicity, their retinal degeneration and viability were largely rescued. Based on these data, we suggest that nuclear accumulation of Staufen is an important feature of C9-ALS/FTD and suggest that reducing stau dosage is a promising therapeutic target.

Multiscale wrinkled structures are widely used in practical applications, such as smart adhesion, surface engineering, and stretchable electronics owing to their easy and low-cost strategy for controlling surface morphologies. This study demonstrates a facile technique for creating multiscale wrinkles of fluorocarbon thin films deposited on soft and stiff polydimethylsiloxane (PDMS) substrates by regulating the elastic properties of the substrates. The elastic properties of the substrates were changed by varying the crosslinking temperatures and the amount of crosslinking agent. The wrinkling structures were strongly influenced by the elastic properties of the substrate during a fluorocarbon polymer sputtering process. For the stiff PDMS substrates, wrinkles with periodic and sinusoidal patterns were observed due to the thermal compression of the bilayer system after deposition. Spontaneous hierarchical structures were generated on soft PDMS substrates by the penetration of fluorocarbon molecules into the PDMS substrates during the sputtering process.

A new brown button mushroom cultivar, ‘Hogam’, C34 line, was made by crossing homokaryons, ASI1164-37 andASI1175-66, selected by RAPD analysis and by cultivating three times. Mycelium of ‘Hogam’ on CDA (compost dextrose agar)media grew well at 25 o C. The optimum pin-heading temperature of new variety and optimum growing temperature was 14-18 o C. The thickness of the mature cap and stipe were thicker than a control, ‘Dahyang’ that developed in 2010. The color ofpileus was light brown and lighter than ‘Dahyang’. Days required from casing to first harvesting were three days longer thancontrol strain, but the weight of harvested fruiting body increased by 1.35 times. ‘Hogam‘ cultivar are expected to contribute tothe diversification of domestic mushroom cultivars.KEYWORDS: Agaricus bisporus, Brown button mushroom cultivar, Morphological and cultural characteristics 서론 양송이(Agaricus bisporus)는 생긴 모양을 빗대어 buttonmushroom으로 불리며, 식용버섯으로 맛과 향기가 뛰어나서 세계적으로 가장 널리 소비되는 버섯 중 하나이다(Jzsefet al. 2004). 최근 우리나라의 양송이 생산량은 6,678MT이고, 생산액은 433억이다(Special crop production in 2013).양송이에는 일반적으로 많이 소비되는 백색양송이 와 갓이갈색을 띄는 갈색양송이로 구성된다. 특히 갈색양송이는크래미버섯, 베이비 포토벨로, 베이비벨라, 미니벨라, 포터벨리니, 로먼버섯, 이탈리아버섯 또는 갈색버섯등과 같은여러 이름으로 불리며 일반 버섯보다 크게 키워, 갓이 약간벌어진 형태의 버섯은 potobello라고 불린다. 우리나라에서는 갈색양송이는 병충해에 강해 과거 705 등을 재배하였으나 가격이 낮고 재배규모는 적어 학교급식을 납품하는 농가에서 소규모로 재배를 하고 있다.양송이는 다른 버섯과 달리 이차 자웅동주성(secondaryhomothalism)으로 한 담포자내에 두 개의 반수체핵(haploidnuclei) 을가지는다핵성(multinucleate) 지닌다. 이에따라 교잡을 하지 않아도 포자내에 화합성을 가진 두 개의 핵이 임성을 띄어 대다수의 담자포자가 자실체를 형성한다.이에교잡을위한동핵균주확보가어렵고, 교잡후꺽쇠연결체(clamp connection)가 형성되지 않아 교잡을 확인하기가 어렵다(Rapar and Raper, 1972). 현재까지 육종연구는돌연변이(Fritsche, 1965; Stoller and Stauffer, 1953), 교배(Elliott, 1978; Kneebone, 1972; Moessner, 1962), 원형질체분리(Horgen, 1991), 형질전환(Rhee et al, 1996; Chen etal, 2000; Tomas et al, 2001) 등이 이루어졌다. 동핵균주의

초피와 번행으로부터 항균성분 추출방법 및 항균조사분쇄한 초피 100 g을 1L 라운드 플라스크에 넣고 증류수 약 500 mL를 넣어 하루 동안 교반한다. 증류수를 걸러낸후 초피를 건져내어 1L 라운드 플라스크에 넣고 hexane500 mL를 넣어 다시 하루 동안 교반한다. Hexane층을 분리한 후 걸러낸 초피를 다시 1L 라운드 플라스크에 넣고ethylacetate 500 mL를 넣고 2일동안 교반 한 후 초피를 걸러내고 ethylacetate층 용액을 모아서 rotatory evaporator를이용하여 농축시킨다. 이 용액을 silicagel을 이용하여 여과시킨 후 불순물을 제거하고 다시 농축시킨다. 얻어진 초피추출액을 초피 I이라 한다.잘게 부순 번행 1kg 을 플라스크에 넣고 methanol 1 L를넣어 3일 정도 교반 한 다음 methanol을 분리한 후 rotatoryevaporator를 이용하여 농축시킨다. 이 농축액을 번행I이라 한다. 번행I을 Kugelroh를 이용하여 감압하고 온도 별로증류한다. 72−76 C 사이에서 나온 증류 액을 번행II라하고, 80−82

Carbonate-type macrodiols synthesized by base-catalyzed polycondensation of co-diols and dimethyl carbonate as an environmentally-friendly route were subsequently utilized for the preparation of transparent and self-healable thermoplastic polyurethanes (TPUs) containing a carbonate-type soft segment. Three types of macrodiols, obtained from mono, dual and triple diol-monomers for target molecular weights of 1 and 1.5 kg mol-1, were analyzed by ¹H NMR integration and the OH titration value. Colorless transparent macrodiols in a liquid state at a room temperature of 20 °C were obtained, except the macrodiol from mono 1,6-hexanediol. Before TPU synthesis, macrodiols require pH neutralization to prevent gelation. TPUs synthesized by a solution pre-polymer method with 4,4'-methylene(bisphenyl isocyanate) and 1,4-butanediol as a chain extender exhibited moderate molecular weights, good transparencies and robust mechanical properties. Especially, the incorporation of 3-methyl-1,5-pentanediol within carbonate-type macrodiols enhanced the transparency of the resultant TPUs by decreasing the degree of microphase separation evidenced by ATR-FTIR and DSC. Interestingly, packing density of hard segments and the degree of microphase separation determined the self-healing efficiency of TPUs, which showed good performances in the case of sourced macrodiols from triple diol-monomers.

Bacterial and fungal pathogens have caused serious problems to the human health. This is particularly true for untreatable infectious diseases and clinical situations where there is no reliable treatment for infected patients. To increase the antimicrobial activity of materials, we introduce silver nanoparticle (NP) patches in which the NPs are incorporated to the surface of smooth and uniform poly(acrylic acid) (PAA) nanofibers. The PAA nanofibers were thermally crosslinked with ethylene glycol via heat treatment through a mild method. The characterization of the resulting PAA-silver NP patches was done using scanning electron microscopy (SEM), UV spectroscopy, X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS). To demonstrate the antimicrobial activity of PAA, we incorporated the patches containing the silver NPs into strains of fungi such as Candida albicans (C. albican) and bacteria such as Methicillin-resistant Staphylococcus aureus (MRSA). The PAA-silver fibers achieved zones of inhibition against C. albicans and MRSA indicating their antimicrobial activity against both fungi and bacteria. We conclude that silver NP patches exhibited multiple inhibitory actions for the interruption and blockage of activity fungal and bacterial strains, which has the potential as an antimicrobial agent in infectious diseases. Moreover, the proposed material has the potential to be used in antimicrobial textile fabrics, food packaging films, and wound dressings.

Changes in fermentation properties and ornithine levels of Baechu Kimchi by storage conditions were investigated. After making and fermenting Kimchi at 15℃ for 32 hr (S1), 36 hr (S2), 40 hr (S3), 44 hr (S4), and 48 hr (S5) during the first 10 days of storage. The Kimchi samples are subsequently stored in the 1℃ Kimchi refrigerator for up to 60 days. Changes in the pH values and lactic acid contents of S4 and S5 samples are slightly bigger than the S1, S2 and S3 samples which have no significance differences. According to lactic acid bacteria (LAB) number, all samples show the largest augmentation according to the number of Lactobacilli during the first 20 days of storage. After 20 days of storage, the S4 and S5 samples show larger accumulations of LAB than S1, S2 and S3 samples. The Weissella genus is predominated at the 40 day of storage in the S5 sample. Ornithine levels are increased up to 170 ㎎ per 100 g during the storage period of 40~50 days in the S5 sample. However, the increase of ornithine levels in S1, S2 and S3 samples is smaller than those of the S4 and S5 samples. These results indicate that the conditions of Kimchi fermentation, which is 48 hr at 15℃ before storage, is proved to be the most superior for ornithine levels within the Kimchi refrigerator. Key words: Kimchi, fermentation, storage, ornithine levels, Weissella genus

Tetraacetylethylenediamine (TAED) was prepared by acetylation of diacetylethylenediamine (DAED) with acetic anhydride in a 5 L reactor coupled with a packed distillation column, 2.5 cm in inside di...

Biodegradable polylactide (PLA) nanofibers containing silver nanoparticles were prepared by electrospinning. Solutions of PLA in dichloromethane/N,N-dimethylacetamide (DMAc) with different amounts of silver nitrate (AgNO3) were used to spin nanofibers. DMAc was used as a reducing agent for Ag+ ions in the PLA solution. In the dilute solution system, the intrinsic viscosity and Huggins constant of the PLA/Ag solutions decreased at AgNO3 concentrations < 6 wt% whereas they increased at 9 wt% AgNO3. On the other hand, dichloromethane/DMAc was found to be a good solvent with a similar solvating power to the Huggins constant results. The solution properties of the viscosity and electrical conductivity of the spinning solutions are strongly affected by the addition of AgNO3. The mean diameters of the PLA/Ag nanofibers containing 0, 3, 6 and 9 wt% of AgNO3 were 400.9±0.1, 352.6±0.3, 313.1±0.2 and 482.8±0.2 nm, respectively. The silver nanoparticles contributed to the formation of thinner PLA/Ag nanofibers. When the PLA nanofibers contained 6 wt% AgNO3, silver nanoparticles with an average size of 4.9 nm were distributed homogeneously in the PLA nanaofibers.

Novel disubstituted adamantyl derivatives were synthesized and evaluated in a P-glycoprotein dependent multidrug resistance cancer cell line. The hit to lead optimization provided potent MDR reversal agents. Some potent adamantyl derivatives were more than 10-fold more potent than verapamil without considerable intrinsic cytotoxicity. The 3-trifluorophenyl derivative 14f did not affect the metabolism of CYP450 3A4, whereas most of MDR revertants had a weak inhibitory effect.

The effects of pretreatment reagents on the hydrolysis and physical properties of PET fabrics were investigated under various alkaline hydrolysis treatment and pretreatment conditions. Before alkaline hydrolysis, solvent treatment with pretreatment reagents, including benzyl alcohol (PET-b) or 2-phenyl ethanol (PET-p), modified the structure of the PET fabric, thus affecting the hydrolysis and physical properties of the PET fabrics. Fabric weight loss increased with increasing hydrolysis time. The rate constant (k) increased markedly with increasing methyl groups in the pretreatment reagents. The activation energy (Ea) of untreated fabrics was higher than those of the treated fabrics. The crystallinity of the PET fabrics increased with increasing hydrolysis times (t) and methyl groups in the pretreatment reagents. The weight loss of PET-b increased with increasing pretreatment temperature (T). However, the weight loss of PET-p increased up to 100°C but decreased above 120°C. The shrinkage of all samples increased with increasing hydrolysis times (t). Shrinkage of PET-b and PET-p was greater than that of untreated fabrics. PET-b displayed greater shrinkage than PET-p because byproducts polluted the PET fibers. Both the initial and maximum water absorption of the fabrics increased with increasing hydrolysis times (t). © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009

Tumor necrosis factor-alpha (TNF-alpha) induces the activation of all three types of mitogen-activated protein kinase (MAPK): c-Jun NH(2)-terminal kinase (JNK), p38, and extracellular signal-regulated kinase (ERK). This cytokine also induces the production of several types of reactive oxygen species, including H(2)O(2). With the use both of HeLa cells expressing wild-type or dominant negative forms of the cytosolic peroxidase peroxiredoxin II and of mouse embryonic fibroblasts deficient in this protein, we evaluated the roles of H(2)O(2) in the activation of MAPKs by TNF-alpha. In vitro kinase assays as well as immunoblot analysis with antibodies specific for activated MAPKs indicated that H(2)O(2) produced in response to TNF-alpha potentiates the activation of JNK and p38 induced by this cytokine but inhibits that of ERK. Our results also suggest that cytosolic peroxiredoxins are important regulators of TNF signaling pathways.

Lipoxygenases (LOXs) catalyze the formation of fatty acid hydroperoxides involved in responses to stresses. This study examines the expression of a non-traditional dual positional specific maize LOX in response to wounding or methyl jasmonate (MeJA). Full-length maize LOX cDNA was expressed in Escherichia coli, and recombinant LOX was purified and characterized enzymatically. RP-HPLC and GC-MS analysis showed that the purified LOX converts alpha-linolenic acid into 13-hydroperoxylinolenic acid and 9-hydroperoxylinolenic acid in a 6:4 ratio. LOX mRNA accumulated rapidly and transiently in response to wounding reaching a peak of expression about 3 h after wounding. This increase followed an initial increase in endogenous jasmonic acid (JA) 1 h after wounding (JA burst). However, the expression of LOX induced by MeJA lasted longer than the expression induced by wounding, and the MeJA-induced expression seemed to be biphasic pattern composed of early and late phases. The expression of LOX in the presence of inhibitors of JA biosynthesis was not completely inhibited, but delayed in wound response and the expression period was shortened in MeJA response. These results suggest that wound-responsive JA burst may trigger the early phase of LOX expression which facilitates biosynthesis of endogenous JA through its 13-LOX activity, and subsequently leads to the activation of the late phase LOX expression in MeJA-treated maize seedlings. Implications of dual positional specificity of maize LOX in the observed expression kinetics are discussed.

A new gene encoding an α-amylase has been cloned, sequenced and expressed in E. coli from an alkaliphilic Pseudomonas sp. KFCC10818. The structural gene is 1356 base pairs long and encodes a protein of 452 amino acids. The recombinant α-amylase has been purified and biochemically characterized. Molecular mass of the protein deduced from SDS-PAGE was 50 kDa. The enzyme showed an activity optimum at pH 8 and at 40 °C with complete stability at pH 13 for 3 h. The enzyme released maltose and maltotriose on hydrolysis of soluble starch. Amylose was hydrolysed over 5 times faster than amylopectin by the enzyme while the hydrolysis of cyclodextrin or pullulan was negligible.

A gene coding for amylase was cloned and sequenced from an alkalophilic Pseudomonas sp. KFCC 10818. The coding region for the amylase precursor contained 1,692 nucleotides. The presumed Shine-Dalgarno sequence, AAGG, was located at 8 nucleotides upstream from the ATG initiation codon. The precursor protein had a putative signal peptide of 25 amino acid residues at its amino terminus. The Pseudomonas amylase had four highly conserved regions as other α-amylases. The amylase expressed from E. coli harboring the Pseudomonas gene produced maltose and maltotriose from soluble starch.

The overexpression of P-glycoprotein (P-gp), an ATP-binding cassette (ABC) transporter, has been demonstrated not only to be primarily involved in the incapacitation of a variety of widely used anticancer agents, such as, paclitaxel (Taxol), doxorubicin, and vinorelbine, but also is observed much more frequently than other ABC transporters. However, despite the developments of a number of P-gp inhibitors to overcome MDR, no drug is currently clinically available. A number of drug candidates have been dropped because of critical limitations, such as, intrinsic toxicity and pharmacokinetic interactions. Furthermore, few accurate methods are available for identifying patients that might benefit from MDR inhibition. However, advanced functional imaging technology using

We conducted a genetic yeast screen to identify salt tolerance (SAT) genes in a maize kernel cDNA library. During the screening, we identified a maize clone (SAT41) that seemed to confer elevated salt tolerance in comparison to control cells. SAT41 cDNA encodes a 16-kDa protein which is 82.4% identical to the Arabidopsis Multiprotein bridging factor 1a (MBF1a) transcriptional coactivator gene. To further examine salinity tolerance in Arabidopsis, we functionally characterized the MBF1a gene and found that dehydration as well as heightened glucose (Glc) induced MBF1a expression. Constitutive expression of MBF1a in Arabidopsis led to elevated salt tolerance in transgenic lines. Interestingly, plants overexpressing MBF1a exhibited insensitivity to Glc and resistance to fungal disease. Our results suggest that MBF1a is involved in stress tolerance as well as in ethylene and Glc signaling in Arabidopsis.

In an effort to asses the effect of Val311Met point mutation of Bacillus subtilis protoporphyrinogen oxidase on the resistance to diphenyl ether herbicides, a Val311Met point mutant of B. subtilis protoporphyrinogen oxidase was prepared, heterologously expressed in Escherichia coli, and the purified recombinant Val311Met mutant protoporphyrinogen oxidase was kinetically characterized. The mutant protoporphyrinogen oxidase showed very similar kinetic patterns to wild type protoporphyrinogen oxidase, with slightly decreased activity dependent on pH and the concentrations of NaCl, Tween 20, and imidazole. When oxyfluorfen was used as a competitive inhibitor, the Val311Met mutant protoporphyrinogen oxidase showed an increased inhibition constant about 1.5 times that of wild type protoporphyrinogen oxidase. The marginal increase of the inhibition constant indicates that the Val311Met point mutation in B. subtilis protoporphyrinogen oxidase may not be an important determinant in the mechanism that protects protoporphyrinogen oxidase against diphenyl ether herbicides.

The overexpression of P-glycoprotein (P-gp), an ATP-binding cassette (ABC) transporter, has been demonstrated not only to be primarily involved in the incapacitation of a variety of widely used anticancer agents, such as, paclitaxel (Taxol), doxorubicin, and vinorelbine, but also is observed much more frequently than other ABC transporters. However, despite the developments of a number of P-gp inhibitors to overcome MDR, no drug is currently clinically available. A number of drug candidates have been dropped because of critical limitations, such as, intrinsic toxicity and pharmacokinetic interactions. Furthermore, few accurate methods are available for identifying patients that might benefit from MDR inhibition. However, advanced functional imaging technology using