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The rumen bacterium Cellulosilyticum ruminicola H1 efficiently hydrolyzes cellulose. To gain insights into the regulatory mechanisms of cellulase synthesis, comparative transcriptome analysis was conducted for cultures grown on 2% filter paper, 0.5 and 0.05% cellobiose, and 0.5% birchwood xylan. It was found that cellulose induced a majority of (hemi)cellulases, including 33 cellulases and a cellulosomal scaffoldin (1.3- to 22.7-fold); seven endoxylanases, two mannanases, and two pectatelyases (2- to 16- fold); and pyruvate formate-lyase (PFL, 1.5- to 7-fold). Noticeably, 3- and 2.5-fold increased transcription of a cellobiohydrolase and the cellulosomal scaffoldin precursor were detected in 0.05% than in 0.5% cellobiose. Consistently, 9- and 4-fold higher specific cellobiohydrolase activities were detected in the filter paper and 0.05%cellobiose culture. SDS- and native-PAGE zymograms of cellulose-enriched proteins from the filter paper culture displayed cellulase activities, and cellulolytic "complexes" were enriched from the filter paper- and 0.05% cellobiose-cultures, but not from the 0.5% cellobiose culture. LC-MS/MS identified the cellulosomal scaffoldin precursor in the "complexes" in addition to cellulase, hemicellulase, and PFL proteins. The addition of 0.5% cellobiose, but not 0.05%cellobiose remarkably inhibited strain H1 to degrade filter paper. Therefore, this work reveals a cellobiose-dose related regulatory mechanism of cellulase synthesis by lower for induction and higher for repression, which has extended our understanding of the regulation of microbial cellulase synthesis. [ABSTRACT FROM AUTHOR]

We tested the hypothesis that elastic binding of the abdomen (AB) would enhance neuromuscular efficiency of the human diaphragm during exercise. Twelve healthy non-obese men aged 24.8 ± 1.7 years (mean ± SE) completed a symptom-limited constant-load cycle endurance exercise test at 85% of their peak incremental power output with diaphragmatic electromyography (EMGdi) and respiratory pressure measurements under two randomly assigned conditions: unbound control (CTRL) and AB sufficient to increase end-expiratory gastric pressure (Pga,ee) by 5-8 cmH2O at rest. By design, AB increased Pga,ee by 6.6 ± 0.6 cmH2O at rest. Compared to CTRL, AB significantly increased the transdiaphragmatic pressure swing-to-EMGdi ratio by 85-95%during exercise, reflecting enhanced neuromuscular efficiency of the diaphragm. By contrast, AB had no effect on spirometric parameters at rest, exercise endurance time or an effect on cardiac, metabolic, ventilatory, breathing pattern, dynamic operating lung volume, and perceptual responses during exercise. In conclusion, AB was associated with isolated and acute improvements in neuromuscular efficiency of the diaphragm during exercise in healthy men. The implications of our results are that AB may be an effective means of enhancing neuromuscular efficiency of the diaphragm in clinical populations with diaphragmatic weakness/dysfunction. [ABSTRACT FROM AUTHOR]

Introduction: Obstructive sleep apnoea (OSA) is characterised by a loss of neuromuscular tone of the upper airway dilator muscles while asleep. This study investigated the effectiveness of transcutaneous electrical stimulation in patients with OSA.Patients and Methods: This was a randomised, sham-controlled crossover trial using transcutaneous electrical stimulation of the upper airway dilator muscles in patients with confirmed OSA. Patients were randomly assigned to one night of sham stimulation and one night of active treatment. The primary outcome was the 4% oxygen desaturation index, responders were defined as patients with a reduction >25% in the oxygen desaturation index when compared with sham stimulation and/or with an index <5/hour in the active treatment night.Results: In 36 patients (age mean 50.8 (SD 11.2) years, male/female 30/6, body mass index median 29.6 (IQR 26.9-34.9) kg/m(2), Epworth Sleepiness Scale 10.5 (4.6) points, oxygen desaturation index median 25.7 (16.0-49.1)/hour, apnoea-hypopnoea index median 28.1 (19.0-57.0)/hour) the primary outcome measure improved when comparing sham stimulation (median 26.9 (17.5-39.5)/hour) with active treatment (median 19.5 (11.6-40.0)/hour; p=0.026), a modest reduction of the mean by 4.1 (95% CI -0.6 to 8.9)/hour. Secondary outcome parameters of patients' perception indicated that stimulation was well tolerated. Responders (47.2%) were predominantly from the mild-to-moderate OSA category. In this subgroup, the oxygen desaturation index was reduced by 10.0 (95% CI 3.9 to 16.0)/hour (p<0.001) and the apnoea-hypopnoea index was reduced by 9.1 (95% CI 2.0 to 16.2)/hour (p=0.004).Conclusion: Transcutaneous electrical stimulation of the pharyngeal dilators during a single night in patients with OSA improves upper airway obstruction and is well tolerated.Trial Registration Number: NCT01661712. [ABSTRACT FROM AUTHOR]

Haloarcula hispanica is one of members of the Halobacteriaceae, which displays particularly low restriction activity and is therefore important as one of the most tractable haloarchaea for archaeal genetic research. Although the Har. hispanica S-layer protein has been reported glycosylated, the S-layer glycoprotein and its glycosylation have not been investigated yet. In this study, the S-layer proteins of Har. hispanica were extracted and characterized. The S-layer was found containing two different glycoproteins which shared highly similar amino acid sequences. The genes coding for these two S-layer glycoproteins were found next to each other in the genome. Moreover, the N- and O-linked glycans were released from these two S-layer glycoproteins for structural determination. Based on the mass spectrometry and nuclear magnetic resonance, the N-glycan was determined as a branched trisaccharide containing a 225 Da residue corresponded to a 2-amino-6-sulfo-2, 6-dideoxy-quinovose, which was the first time that a naturally occurring form of sulfoquinovosamine was identified. Besides, the O-glycan was characterized as a Glca-1,4-Gal disaccharide by mass spectrometry combined with monosaccharide composition analysis and glycosidase treatment. The determination of the N- and O-glycan structure will be helpful for studying the diverse protein glycosylation pathways in archaea utilizing H. hispanica as a new model. [ABSTRACT FROM AUTHOR]

Background: Clostridium thermocellum is a Gram-positive thermophilic anaerobic bacterium with the unusual capacity to convert cellulosic biomass into ethanol and hydrogen. Identification and characterization of protein complexes in C. thermocellum are important toward understanding its metabolism and physiology. Results: A two dimensional blue native/SDS-PAGE procedure was developed to separate membrane protein complexes of C. thermocellum. Proteins spots were identified by MALDI-TOF/TOF Mass spectrometry. 24 proteins were identified representing 13 distinct protein complexes, including several putative intact complexes. Interestingly, subunits of both the F1-F0-ATP synthase and the V1-V0-ATP synthase were detected in the membrane sample, indicating C. thermocellum may use alternative mechanisms for ATP generation. Conclusion: Two dimensional blue native/SDS-PAGE was used to detect membrane protein complexes in C. thermocellum. More than a dozen putative protein complexes were identified, revealing the simultaneous expression of two sets of ATP synthase. The protocol developed in this work paves the way for further functional characterization of these protein complexes. [ABSTRACT FROM AUTHOR]

Background: Senescence-accelerated mice (SAM) are commonly used as animal models for studying aging. To date, aging-related proteomes in the livers and brains of SAMP8 mice have been reported; however, spleen of SAM has not yet been used as material for studying the aging-related proteome despite its extremely important role in the progress of aging or aging-related diseases. Objective: To identify proteins associated with aging or aging-related diseases in SAM. Methods: Two-dimensional gel electrophoresis (2DE) was used to compare the proteomes of splenic lymphocytes from two strains of SAM (SAMP8 and SAMR1). Differentially expressed spots whose expression altered over twofold (p < 0.01) were identified by LC-MS/MS and database search. Results: Identification results showed that 18 differentially expressed protein spots represented 14 different proteins known to be involved in aging- or aging disease-related pathways, i.e., annexin I, calgranulin B, transaldolase, isocitrate dehydrogenase, and an unnamed protein product were involved in oxidative stress and cell defense, phosphoglycerate mutase I, aldose reductase, carbonic anhydrase I and carbonic anhydrase II were involved in glycolytic energy metabolism and homeostasis, and macrophage capping protein and high mobility group box-1 protein were involved in infection and DNA damage. Conclusion: 2DE combined with MS is effective for comparative proteome analysis of splenic lymphocytes of SAM to identify proteins closely involved in aging or aging-related diseases, especially immune diseases, and further functional studies of these proteins in SAM may throw some light on the study of biomarkers for aging or aging-related diseases. Copyright © 2009 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

In yeast, glycosylphosphatidylinositol (GPI) is essential for viability and plays an important role in biosynthesis and organization of cell wall. Initiation of the GPI anchor biosynthesis is catalysed by the GPI– N-acetylglucosaminyltransferase complex (GPI–GnT). The GPI3 ( SPT14) gene is thought to encode the catalytic subunit of GPI–GnT complex. In contrast to Saccharomyces cerevisiae, little is known about the GPI biosynthesis in filamentous fungi. In this study, the afpig-a gene was identified as the homologue of the GPI3/ pig-A gene in Aspergillus fumigatus, an opportunistic fungal pathogen. By replacement of the afpig-a gene with a pyrG gene, we obtained the null mutants. Although the Δ afpig-a mutant exhibited a significant increased cell lysis instead of temperature-sensitive or conditional lethal phenotype associated to the GPI3 mutant of yeast, they could survive at temperatures from 30°C to 50°C. The analysis of the mutants showed that a completely blocking of the GPI anchor synthesis in A. fumigatus led to cell wall defect, abnormal hyphal growth, rapid conidial germination and aberrant conidiation. In vivo assays revealed that the mutant exhibited a reduced virulence in immunocompromised mice. The GPI anchor was not essential for viability, but required for the cell wall integrity, morphogenesis and virulence in A. fumigatus. [ABSTRACT FROM AUTHOR]

The mammalian ER/cytosolic α-mannosidase (Man2C1p), yeast vacuolar α-mannosidase (Ams1p) and the Aspergillus nidulans α-mannosidase are members of Class IIC subgroup, which is involved in oligosaccharide catabolism and N-glycan processing. Unlike their mammalian counterparts, the yeast Ams1p and A. nidulans Class IIC α-mannosidase are not essential for morphogenesis and cellular function. In this study, the Afams1, a gene encoding a member of Class IIC α-mannosidases, was identified in the opportunistic pathogen Aspergillus fumigatus. Deletion of the Afams1 led to a severe defect in conidial formation, especially at a higher temperature. In addition, abnormalities of polarity and septation were associated with the ΔAfams1 mutant. Our results showed that the Afams1 gene, in contrast to its homolog in yeast or A. nidulans, was required for morphogenesis and cellular function in A. fumigatus. [ABSTRACT FROM AUTHOR]