scholarly journals Effects of Altered Fructose 2,6-Bisphosphate Levels on Carbohydrate Metabolism in Carnation

HortScience ◽  
2007 ◽  
Vol 42 (2) ◽  
pp. 403-406 ◽  
Author(s):  
Antal Szőke ◽  
Erzsébet Kiss ◽  
László Heszky ◽  
Ildikó Kerepesi ◽  
Ottó Toldi
Keyword(s):  
Carbohydrate Metabolism ◽  
Dianthus Caryophyllus ◽  
Bifunctional Enzyme ◽  
Starch Metabolism ◽  
Wild Type ◽  
Type Transformation ◽  
Key Enzyme ◽  
Low Levels ◽  
Hexose Phosphates

The aim of this work was to examine the role of fructose 2,6-bisphosphate (fru 2,6P2) in the carbohydrate metabolism in carnation (Dianthus caryophyllus L.). For this purpose, transgenic plants harboring two modified bifunctional enzyme complementary DNAs of rat liver origin (6-phosphofructo-2-kinase/fructose 2,6-biphosphatase) were generated. Transformation with the kinase construct resulted in a 45% to 85% increase in fru 2,6P2 concentrations compared with the wild type. Transformation with the phosphatase construct reduced the fru 2,6P2 contents by 45% and 70%. These alterations in fru 2,6P2 amounts affected the key enzyme activities of sucrose and starch metabolism. Accordingly, plants with elevated fru 2,6P2 concentrations had high levels of starch, fructose, and triose phosphates, and low levels of sucrose, glucose, and hexose phosphates. In plants with reduced amounts of fru 2,6P2 different results could be observed in major carbohydrate compounds.

scholarly journals Platelet and Erythrocyte Extravasation across Inflamed Corneal Venules Depend on CD18, Neutrophils and Mast Cell Degranulation

2021 ◽  
Vol 22 (14) ◽  
pp. 7360
Author(s):  
Angie De La Cruz ◽  
Aubrey Hargrave ◽  
Sri Magadi ◽  
Justin A. Courson ◽  
Paul T. Landry ◽  
...  
Keyword(s):  
Mast Cell ◽  
Cell Types ◽  
Mast Cell Degranulation ◽  
Wild Type ◽  
Delayed Wound Healing ◽  
Corneal Epithelial ◽  
Corneal Abrasion ◽  
Low Levels ◽  
Endothelial Pores

Platelet extravasation during inflammation is under-appreciated. In wild-type (WT) mice, a central corneal epithelial abrasion initiates neutrophil (PMN) and platelet extravasation from peripheral limbal venules. The same injury in mice expressing low levels of the β2-integrin, CD18 (CD18hypo mice) shows reduced platelet extravasation with PMN extravasation apparently unaffected. To better define the role of CD18 on platelet extravasation, we focused on two relevant cell types expressing CD18: PMNs and mast cells. Following corneal abrasion in WT mice, we observed not only extravasated PMNs and platelets but also extravasated erythrocytes (RBCs). Ultrastructural observations of engorged limbal venules showed platelets and RBCs passing through endothelial pores. In contrast, injured CD18hypo mice showed significantly less venule engorgement and markedly reduced platelet and RBC extravasation; mast cell degranulation was also reduced compared to WT mice. Corneal abrasion in mast cell-deficient (KitW-sh/W-sh) mice showed less venule engorgement, delayed PMN extravasation, reduced platelet and RBC extravasation and delayed wound healing compared to WT mice. Finally, antibody-induced depletion of circulating PMNs prior to corneal abrasion reduced mast cell degranulation, venule engorgement, and extravasation of PMNs, platelets, and RBCs. In summary, in the injured cornea, platelet and RBC extravasation depends on CD18, PMNs, and mast cell degranulation.

Role of eNOS-derived NO in the postischemic anti-inflammatory effects of antecedent ethanol ingestion in murine small intestine

2007 ◽  
Vol 292 (3) ◽  
pp. H1435-H1442 ◽  
Author(s):  
Taiji Yamaguchi ◽  
Kazuhiro Kamada ◽  
Catherine Dayton ◽  
F. Spencer Gaskin ◽  
Mozow Yusof ◽  
...  
Keyword(s):  
Ethanol Exposure ◽  
Ethanol Ingestion ◽  
Leukocyte Rolling ◽  
Wild Type ◽  
No Donor ◽  
Nos Inhibitors ◽  
Low Levels ◽  
Ethanol Preconditioning ◽  
Dependent Mechanism

Ingestion of low levels of ethanol 24 h before [ethanol preconditioning (EPC)] ischemia and reperfusion (I/R) prevents postischemic leukocyte rolling (LR) and adhesion (LA), effects that were abolished by adenosine A2 receptor (ADO-A2R) antagonists or nitric oxide (NO) synthase (NOS) inhibitors. The aims of this study were to determine whether NO derived from endothelial NOS (eNOS) during the period of ethanol exposure triggered entrance into this preconditioned state and whether these events were initiated by an ADO-A2R-dependent mechanism. Ethanol or distilled water vehicle was administered to C57BL/6J [wild type (WT)] or eNOS-deficient (eNOS−/−) mice by gavage. Twenty-four hours later, the superior mesenteric artery was occluded for 45 min. LR and LA were quantified by intravital microscopy after 30 and 60 min of reperfusion. I/R increased LR and LA in WT mice, effects that were abolished by EPC or NO donor preconditioning (NO-PC). NO-PC was not attenuated by coincident administration of an ADO-A2R antagonist. I/R increased LR and LA in eNOS−/− mice to levels comparable with those noted in WT animals. However, EPC only slightly attenuated postischemic LR and LA, whereas NO-PC remained effective as a preconditioning stimulus in eNOS−/− mice. Preconditioning with an ADO-A2R agonist (which we previously demonstrated prevents I/R-induced LR and LA in WT animals) failed to attenuate these postischemic adhesive responses in eNOS−/− mice. Our results indicate that EPC is triggered by NO formed secondary to ADO-A2R-dependent eNOS activation during the period of ethanol exposure 24 h before I/R.

scholarly journals In Helicobacter pylori, LuxS Is a Key Enzyme in Cysteine Provision through a Reverse Transsulfuration Pathway

2010 ◽  
Vol 192 (5) ◽  
pp. 1184-1192 ◽  
Author(s):  
Neil C. Doherty ◽  
Feifei Shen ◽  
Nigel M. Halliday ◽  
David A. Barrett ◽  
Kim R. Hardie ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Wild Type ◽  
Metabolic Role ◽  
Absolute Requirement ◽  
Transsulfuration Pathway ◽  
Cysteine Synthesis ◽  
Key Enzyme ◽  
H Pylori ◽  
Isogenic Strains

ABSTRACT In many bacteria, LuxS functions as a quorum-sensing molecule synthase. However, it also has a second, more central metabolic function in the activated methyl cycle (AMC), which generates the S-adenosylmethionine required by methyltransferases and recycles the product via methionine. Helicobacter pylori lacks an enzyme catalyzing homocysteine-to-methionine conversion, rendering the AMC incomplete and thus making any metabolic role of H. pylori LuxS (LuxSHp) unclear. Interestingly, luxS Hp is located next to genes annotated as cysK Hp and metB Hp, involved in other bacteria in cysteine and methionine metabolism. We showed that isogenic strains carrying mutations in luxS Hp, cysK Hp, and metB Hp could not grow without added cysteine (whereas the wild type could), suggesting roles in cysteine synthesis. Growth of the ΔluxS Hp mutant was restored by homocysteine or cystathionine and growth of the ΔcysK Hp mutant by cystathionine only. The ΔmetB Hp mutant had an absolute requirement for cysteine. Metabolite analyses showed that S-ribosylhomocysteine accumulated in the ΔluxS Hp mutant, homocysteine in the ΔcysK Hp mutant, and cystathionine in the ΔmetB Hp mutant. This suggests that S-ribosylhomocysteine is converted by LuxSHp to homocysteine (as in the classic AMC) and thence by CysKHp to cystathionine and by MetBHp to cysteine. In silico analysis suggested that cysK-metB-luxS were acquired by H. pylori from a Gram-positive source. We conclude that cysK-metB-luxS encode the capacity to generate cysteine from products of the incomplete AMC of H. pylori in a process of reverse transsulfuration. We recommend that the misnamed genes cysK Hp and metB Hp be renamed mccA (methionine-to-cysteine-conversion gene A) and mccB, respectively.

scholarly journals Production of transgenic carnation with a heterologous 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase bifunctional enzyme cDNA

2006 ◽  
Vol 12 (4) ◽  
Author(s):  
A. Szőke ◽  
E. Kiss ◽  
O. Toldi ◽  
L. Heszky
Keyword(s):  
Carbohydrate Metabolism ◽  
Transgene Expression ◽  
Dianthus Caryophyllus ◽  
Basal Medium ◽  
Relative Activity ◽  
Pcr Analysis ◽  
Bifunctional Enzyme ◽  
Specific Primers ◽  
Signal Molecule ◽  
And Control

Transgenic carnations were produced with a modified mammalian bifunctional enzyme cDNA coding 6-phosphofructo-2- kinaseffructose 2,6-bisphosphatase. Relative activity of this enzyme determines the fructose 2,6-bisphosphate (fru 2,6-P2) cytosolic concentration. This metabolite — as a signal molecule — is one of the carbohydrate metabolism regulators. The regenerated Dianthus chinensis and Dianthus caryophyllus shoots were selected on MS basal medium containing 150 mg/1 kanamycin. Transgene integration was proven by PCR analysis with cDNA specific primers followed by Southern hybridization of DNA isolated from selected green shoots, which survived on kanamycin containing medium, so 3 D. chinensis and 20 D. caryophyllus transgenic plants were produced. Transgene expression were examined by RT-PCR. Transformed and control plants were potted in glasshouse to evaluate the effect of modified fru 2,6-P2 on development, growth and carbohydrate metabolism.

scholarly journals α2β1 integrin expression in the tumor microenvironment enhances tumor angiogenesis in a tumor cell–specific manner

Blood ◽  
2008 ◽  
Vol 111 (4) ◽  
pp. 1980-1988 ◽  
Author(s):  
Zhonghua Zhang ◽  
Norma E. Ramirez ◽  
Thomas E. Yankeelov ◽  
Zhengzhi Li ◽  
Laura E. Ford ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Tumor Angiogenesis ◽  
Genetic Alterations ◽  
Wild Type ◽  
B16f10 Melanoma ◽  
B16f10 Cells ◽  
Low Levels ◽  
Α2β1 Integrin ◽  
Integrin Α2

To define the role of the α2β1 integrin in pathologic angiogenesis, we investigated tumor-associated growth and angiogenesis in wild-type and α2-null mice. Our findings reveal that the α2β1 integrin plays an important role in angiogenesis via regulation of VEGFR1 expression. When challenged with B16F10 melanoma cells, mice lacking α2β1 integrin ex-pression exhibit increased tumor angiogenesis associated with up-regulated VEGFR1 expression. In contrast, there was no α2β1 integrin-dependent difference in the angiogenic response to Lewis lung carcinoma (LLC) cells. Interestingly, whereas B16F10 cells secrete high levels of placental growth factor (PLGF), LLC cells produce high levels of VEGF, but low levels of PLGF. The α2β1 integrin-dependent difference in angiogenesis was restored to LLC cells by expression of PLGF, strongly suggesting that the angiogenic phenotype and tumor growth in the α2-null host is dependent on specific interactions between the tumor cell and the genetically defined integrin repertoire of the host microenvironment. Thus integrin α2-null mice represent an example of genetic alterations of “the soil” determining response to the “seed.”

Role of the N-terminus in human 4-hydroxyphenylpyruvate dioxygenase activity

2019 ◽  
Vol 167 (3) ◽  
pp. 315-322
Author(s):  
An-Ning Feng ◽  
Chih-Wei Huang ◽  
Chi-Huei Lin ◽  
Yung-Lung Chang ◽  
Meng-Yuan Ni ◽  
...  
Keyword(s):  
Active Site ◽  
Catalytic Efficiency ◽  
Dynamics Simulation ◽  
Wild Type ◽  
Catalytic Function ◽  
C Terminus ◽  
N Terminus ◽  
Key Enzyme ◽  
The Stability

Abstract 4-Hydroxyphenylpyruvate dioxygenase (HPPD) is a key enzyme in tyrosine catabolism, catalysing the oxidation of 4-hydroxyphenylpyruvate to homogentisate. Genetic deficiency of this enzyme causes type III tyrosinaemia. The enzyme comprises two barrel-shaped domains formed by the N- and C-termini, with the active site located in the C-terminus. This study investigated the role of the N-terminus, located at the domain interface, in HPPD activity. We observed that the kcat/Km decreased ∼8-fold compared with wild type upon removal of the 12 N-terminal residues (ΔR13). Interestingly, the wild-type level of activity was retained in a mutant missing the 17 N-terminal residues, with a kcat/Km 11-fold higher than that of the ΔR13 mutant; however, the structural stability of this mutant was lower than that of wild type. A 2-fold decrease in catalytic efficiency was observed for the K10A and E12A mutants, indicating synergism between these residues in the enzyme catalytic function. A molecular dynamics simulation showed large RMS fluctuations in ΔR13 suggesting that conformational flexibility at the domain interface leads to lower activity in this mutant. These results demonstrate that the N-terminus maintains the stability of the domain interface to allow for catalysis at the active site of HPPD.

scholarly journals Role of Phosphoglucomutase of Stenotrophomonasmaltophilia in Lipopolysaccharide Biosynthesis, Virulence, and Antibiotic Resistance

2003 ◽  
Vol 71 (6) ◽  
pp. 3068-3075 ◽  
Author(s):  
Geoffrey A. McKay ◽  
Donald E. Woods ◽  
Kelly L. MacDonald ◽  
Keith Poole
Keyword(s):  
Animal Model ◽  
Antibiotic Resistance ◽  
Parent Strain ◽  
Antimicrobial Agents ◽  
Bifunctional Enzyme ◽  
Wild Type ◽  
Lipopolysaccharide Biosynthesis ◽  
Wild Type Parent ◽  
Alginate Biosynthesis

ABSTRACT A homologue of the algC gene, responsible for the production of a phosphoglucomutase (PGM) associated with LPS and alginate biosynthesis in Pseudomonas aeruginosa, spgM, was cloned from Stenotrophomonas maltophilia. The spgM gene was shown to encode a bifunctional enzyme with both PGM and phosphomannomutase activities. Mutants lacking spgM produced less LPS than the SpgM+ parent strain and had a tendency for shorter O polysaccharide chains. No changes in LPS chemistry were obvious as a result of the loss of spgM. Significantly, however, spgM mutants displayed a modest increase in susceptibility to several antimicrobial agents and were completely avirulent in an animal model of infection. The latter finding may relate to the resultant serum sensitivity of spgM mutants which, unlike the wild-type parent strain, were rapidly killed by human serum. These data highlight the contribution made by LPS to the antimicrobial resistance and virulence of S. maltophilia.

scholarly journals Role of the Haemophilus ducreyi Ton System in Internalization of Heme from Hemoglobin

1998 ◽  
Vol 66 (1) ◽  
pp. 151-160 ◽  
Author(s):  
Christopher Elkins ◽  
Pat A. Totten ◽  
Bonnie Olsen ◽  
Christopher E. Thomas
Keyword(s):  
Outer Membrane Proteins ◽  
Amino Acid Sequences ◽  
Haemophilus Ducreyi ◽  
Wild Type ◽  
E Coli ◽  
Allelic Replacement ◽  
Low Levels ◽  
High Concentrations ◽  
Isogenic Mutants

ABSTRACT By cloning into Escherichia coli and construction of isogenic mutants of Haemophilus ducreyi, we showed that the hemoglobin receptor (HgbA) is TonB dependent. An E. coli hemA tonB mutant expressing H. ducreyi hgbA grew on low levels of hemoglobin as a source of heme only when an intact H. ducreyi Ton system plasmid was present. In contrast, growth on heme by the E. coli hemA tonB mutant expressinghgbA was observed only at high concentrations of heme, was TonB independent, and demonstrated that H. ducreyi HgbA was not sufficient to function as a typical TonB-dependent heme receptor inE. coli. Allelic replacement of the wild-type H. ducreyi exbB, exbD, and tonB loci with the exbB, exbD, and tonB deletion resulted in an H. ducreyi isogenic mutant unable to utilize hemoglobin but able to utilize hemin at the same levels as the parent strain to fulfill its heme requirement. This finding confirms the TonB dependence of HgbA-mediated hemoglobin utilization and suggests that uptake of hemin in H. ducreyi is TonB independent. Additionally, the H. ducreyi Ton system mutant synthesized increased amounts of HgbA and other heme-regulated outer membrane proteins, consistent with derepression of these proteins due to lower intracellular heme and/or iron concentrations in the mutant. Sequencing of the Ton system genes revealed that the arrangement of the genes wasexbB exbD tonB. The proximity and structure of these genes suggested that they are transcribed as an operon. This arrangement, as well as the DNA and deduced amino acid sequences of these H. ducreyi genes, was most similar to those from other pasteurellae.

The role of ICAM-1 in endotoxin-induced acute renal failure

2007 ◽  
Vol 293 (4) ◽  
pp. F1262-F1271 ◽  
Author(s):  
Xiaoyan Wu ◽  
Rongqing Guo ◽  
Ying Wang ◽  
Patrick N. Cunningham
Keyword(s):  
Renal Failure ◽  
Acute Renal Failure ◽  
Adhesion Molecule ◽  
Neutrophil Infiltration ◽  
Intercellular Adhesion Molecule ◽  
Wild Type ◽  
Intercellular Adhesion Molecule 1 ◽  
Low Levels ◽  
Necrosis Factor

The pathogenesis of acute renal failure (ARF) occurring during the course of sepsis is incompletely understood. Intercellular adhesion molecule-1 (ICAM-1) is a key cell adhesion molecule upregulated by LPS, which binds to the integrins CD11a/CD18 and CD11b/CD18 present on the surface of leukocytes. We hypothesized that ICAM-1 facilitates renal injury in LPS-induced ARF. To test this, three groups of mice ( n = 8 per group) were injected intraperitoneally with 6 mg/kg LPS: 1) normal C57BL/6 mice, 2) mice with a targeted deficiency of ICAM-1 (ICAM-1−/−), and 3) mice expressing very low levels of CD18 (CD18-def). ICAM-1−/− mice were significantly resistant to LPS-mediated ARF, as opposed to CD18-def mice, which developed severe ARF, as did wild-type controls (48 h blood urea nitrogen 143 ± 31.5, 70.8 ± 24.4, and 185 ± 16.6 mg/dl in wild-type, ICAM-1−/−, and CD18-def mice, respectively, P < 0.05). At death, ICAM-1−/− mice had significantly less renal neutrophil infiltration than the other two groups, as well as less histological tubular injury. Depletion of neutrophils with mAb Gr-1 led to a profound exaggeration of tumor necrosis factor (TNF) release and high mortality, but neutrophil-depleted mice receiving 10-fold less LPS were protected against ARF despite TNF release similar to what is normally associated with LPS-induced ARF. LPS caused a significant increase in renal expression of chemokines; however, this increase was significantly exaggerated in CD18-def mice, which may account for their lack of protection. In conclusion, these data show that ICAM-1 plays a key role in LPS-induced ARF.

scholarly journals Hsp90 Is Required for Pheromone Signaling in Yeast

1998 ◽  
Vol 9 (11) ◽  
pp. 3071-3083 ◽  
Author(s):  
Jean-François Louvion ◽  
Toufik Abbas-Terki ◽  
Didier Picard
Keyword(s):  
Budding Yeast ◽  
Heat Shock Protein 90 ◽  
Wild Type ◽  
Pheromone Signaling ◽  
Endogenous Substrate ◽  
Low Levels ◽  
Biochemical Analyses ◽  
Map Kinase Pathways

The heat-shock protein 90 (Hsp90) is a cytosolic molecular chaperone that is highly abundant even at normal temperature. Specific functions for Hsp90 have been proposed based on the characterization of its interactions with certain transcription factors and kinases including Raf in vertebrates and flies. We therefore decided to address the role of Hsp90 for MAP kinase pathways in the budding yeast, an organism amenable to both genetic and biochemical analyses. We found that both basal and induced activities of the pheromone-signaling pathway depend on Hsp90. Signaling is defective in strains expressing low levels or point mutants of yeast Hsp90 (Hsp82), or human Hsp90β instead of the wild-type protein. Ste11, a yeast equivalent of Raf, forms complexes with wild-type Hsp90 and depends on Hsp90 function for accumulation. For budding yeast, Ste11 represents the first identified endogenous “substrate” of Hsp90. Moreover, Hsp90 functions in steroid receptor and pheromone signaling can be genetically separated as the Hsp82 point mutant T525I and the human Hsp90β are specifically defective for the former and the latter, respectively. These findings further corroborate the view that molecular chaperones must also be considered as transient or stable components of signal transduction pathways.

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