Functional dissection of alternative secretory pathways in the yeast S. cerevisiae

Eukaryotic cells are characterized by having a subset of internal membrane compartments, each one with a specifi c identity, structure and function. Proteins destined to be targeted to the exterior of the cell need to enter and progress through the secretory pathway. Transport of secretory proteins from the endoplasmic reticulum (ER) to the Golgi takes place by the selective packaging of proteins into COPII-coated vesicles at the ER membrane. Taking advantage of the extensive genetic tools available for S. cerevisiae we found that Hsp150, a yeast secretory glycoprotein, selectively exited the ER in the absence of any of the three Sec24p family members. Sec24p has been thought to be an essential component of the COPII coat and thus indispensable for exocytic membrane traffic. Next we analyzed the ability of Hsp150 to be secreted in mutants, where post-Golgi transport is temperature sensitive. We found that Hsp150 could be selectively secreted under conditions where the exocyst component Sec15p is defective. Analysis of the secretory vesicles revealed that Hsp150 was packaged into a subset of known secretory vesicles as well as in a novel pool of secretory vesicles at the level of the Golgi. Secretion of Hsp150 in the absence of Sec15p function was dependent of Mso1p, a protein capable of interacting with vesicles intended to fuse with the plasma membrane, with the SNARE machinery and with Sec1p. This work demonstrated that Hsp150 is capable of using alternative secretory pathways in ER-to-Golgi and Golgi-to-plasma membrane traffi c. The sorting signals, used at both stages of the secretory pathway, for secretion of Hsp150 were different, revealing the highly dynamic nature and spatial organization of the secretory pathway. Foreign proteins usually misfold in the yeast ER. We used Hsp150 as a carrier to assist folding and transport of heterologous proteins though the secretory pathway to the culture medium in both S. cerevisiae and P. pastoris. Using this technique we expressed Hsp150Δ-HRP and developed a staining procedure, which allowed the visualization of the organelles of the secretory pathway of S. cerevisiae.

Dynamic viscosity of ternary mixtures of dichlorodifluoromethane (R-12), chlorodifluoromethane (R-22), and dichlorotetrafluoroethane (R-114) vapors

The viscosities of ternary mixtures of R-12, R-22, and R-114 vapors were determined at ambient temperature and pressure within +-1% by using an oscillating disk viscometer. The empirical viscosity obtained by Wllke's equation compares very well with the experimental results obtained with this vlscometer. In the case of this ternary vapor mixture, as long as the molar fraction ratio of R-12 to R-114 Is maintained at approximately 2"' (=Inverse ratio of thelr molecular weights) the viscosity of the ternary mixture at ambient temperature and pressure remalns constant irrespective of the percentage of R-22 present in the mixture.

Phospholipids of lipid-containing bacteriophages and their transbilayer distribution

In this study we used electro-spray ionization mass-spectrometry to determine phospholipid class and molecular species compositions in bacteriophages PM2, PRD1, Bam35 and phi6 as well as their hosts. To obtain compositional data of the individual leaflets, phospholipid transbilayer distribution in the viral membranes was studied. We found that 1) the membranes of all studied bacteriophage are enriched in PG as compared to the host membranes, 2) molecular species compositions in the phage and host membranes are similar, and 3) phospholipids in the viral membranes are distributed asymmetrically with phosphatidylglycerol enriched in the outer leaflet and phosphatidylethanolamine in the inner one (except Bam35). Alternative models for selective incorporation of phospholipids to phages and for the origins of the asymmetric phospholipid transbilayer distribution are discussed. Notably, the present data are also useful when constructing high resolution structural models of bacteriophages, since diffraction methods cannot provide a detailed structure of the membrane due to high motility of the lipids and lack of symmetric organization of membrane proteins.

CancerLectinDB: a database of lectins relevant to cancer

The role of lectins in mediating cancer metastasis, apoptosis as well as various other signaling events has been well established in the past few years. Data on various aspects of the role of lectins in cancer is being accumulated at a rapid pace. The data on lectins available in the literature is so diverse, that it becomes difficult and time-consuming, if not impossible to comprehend the advances in various areas and obtain the maximum benefit. Not only do the lectins vary significantly in their individual functional roles, but they are also diverse in their sequences, structures, binding site architectures, quaternary structures, carbohydrate affinities and specificities as well as their potential applications. An organization of these seemingly independent data into a common framework is essential in order to achieve effective use of all the data towards understanding the roles of different lectins in different aspects of cancer and any resulting applications. An integrated knowledge base (CancerLectinDB) together with appropriate analytical tools has therefore been developed for lectins relevant for any aspect of cancer, by collating and integrating diverse data. This database is unique in terms of providing sequence, structural, and functional annotations for lectins from all known sources in cancer and is expected to be a useful addition to the number of glycan related resources now available to the community. The database has been implemented using MySQL on a Linux platform and web-enabled using Perl-CGI and Java tools. Data for individual lectins pertain to taxonomic, biochemical, domain architecture, molecular sequence and structural details as well as carbohydrate specificities. Extensive links have also been provided for relevant bioinformatics resources and analytical tools. Availability of diverse data integrated into a common framework is expected to be of high value for various studies on lectin cancer biology.

Occupational Rhinitis : Diagnosis and Health-related Quality of Life

Occupational rhinitis is mainly caused by work environment and not by stimuli encountered outside the workplace. It differs from rhinitis that is worsened by, but not mainly caused by, workplace exposures. Occupational rhinitis can develop in response to allergens, inhaled irritants, or corrosive gases. The thesis evaluated the use of challenge tests in occupational rhinitis diagnostics, studied the long-term health-related quality of life among allergic occupational rhinitis patients, and the allergens of wheat grain among occupational respiratory allergy patients. The diagnosed occupational rhinitis was mainly allergic rhinitis, which was caused by occupational agents, most commonly flours and animal allergens. The non-IgE-mediated rhinitis reactions were less frequent and caused more often asthma than rhinitis. Both nasal challenges and inhalation challenges were found to be safe tests. The inhalation challenge tests had considerably resource-intensive methodology. However, the evaluation of nasal symptoms and signs together with bronchial reactions saved time and expense compared with the organization of multiple individual challenges. The scoring criteria used matched well with the weighted amount of discharge ≥ 0.2 g and in most cases gave comparable results. The challenge tests are valuable tools when there is uncertainty whether the patient's exposure should be reduced or discontinued. It was found that continuing exposure decreases health-related quality of life among patients with allergic occupational rhinitis despite of rhinitis medications, still approximately ten years after the diagnosis. Health-related quality of life among occupational rhinitis patients without any longer occupational exposure was mainly similar than that of the healthy controls. This highlights the importance of the reduction and cessation of occupational exposure. To achieve this, 17% of occupational rhinitis patients had been re-educated. Alpha-amylase inhibitors, lipid transfer protein 2G, thaumatin -like protein, and peroxidase I were found to be relevant allergens in Finnish patients with occupational respiratory wheat allergy. Of these allergens, thaumatin-like protein and lipid transfer protein 2G were found as new allergens associated with baker's rhinitis and asthma. The knowledge of the new clinically relevant proteins can be used in the future in the development of better standardized diagnostic preparations.

Creep behaviour of nimonic and R.R. 59 alloys

The Ramberg-Osgood relation which adequately describes the stress-strain curve of a strain-hardening material is extended to formulate the constitutive laws for creep. The constitutive laws which describe primary creep adequately are extended to secondary creep. The results are verified for the case of R.R. 59 at 200°C, Nimonic 80A and Nimonic 90 alloys at 750°C.

Root system traits of Norway spruce, Scots pine, and silver birch in mixed boreal forests: an analysis of root architecture, morphology, and anatomy

The aim of this thesis was to unravel the functional-structural characteristics of root systems of Betula pendula Roth., Picea abies (L.) Karst., and Pinus sylvestris L. in mixed boreal forest stands differing in their developmental stage and site fertility. The root systems of these species had similar structural regularities: horizontally-oriented shallow roots defined the horizontal area of influence, and within this area, each species placed fine roots in the uppermost soil layers, while sinker roots defined the maximum rooting depth. Large radial spread and high ramification of coarse roots, and the high specific root length (SRL) and root length density (RLD) of fine roots indicated the high belowground competitiveness and root plasticity of B. pendula. Smaller radial root spread and sparser branching of coarse roots, and low SRL and RLD of fine roots of the conifers could indicate their more conservative resource use and high association with and dependence on ectomycorrhiza-forming fungi. The vertical fine root distributions of the species were mostly overlapping, implying the possibility for intense belowground competition for nutrients. In each species, conduits tapered and their frequency increased from distal roots to the stem, from the stem to the branches, and to leaf petioles in B. pendula. Conduit tapering was organ-specific in each species violating the assumptions of the general vascular scaling model (WBE). This reflects the hierarchical organization of a tree and differences between organs in the relative importance of transport, safety, and mechanical demands. The applied root model was capable of depicting the mass, length and spread of coarse roots of B. pendula and P. abies, and to the lesser extent in P. sylvestris. The roots did not follow self-similar fractal branching, because the parameter values varied within the root systems. Model parameters indicate differences in rooting behavior, and therefore different ecophysiological adaptations between species.

Studies on the metabolism of a monoterpene ketone, R-(+)-pulegone-a hepatotoxin in rat: isolation and characterization of new metabolites

1. The metabolic disposition of R-(+)-pulegone (1) was examined in rats following four daily oral doses (250 mg/kg). 2. Six metabolites, namely pulegol (II), 2-hydroxy-2-(1-hydroxy-1-methylethyl)-5-methylcyclohexanone (III), 3,6-dimethyl-7a-hydroxy-5,6,7,7a-tetrahydro-2(4H)-benzofuranone (IV), menthofuran (V), 5-methyl-2-(1-methyl-1-carboxyethylidene)cyclohexanone (VI), and 5-methyl-5-hydroxy-2-(1-hydroxy-1-carboxyethyl)cyclohexanone (VII) have previously been isolated from rat urine, and identified (Moorthy et al. (1989a). Eight new metabolites have now been isolated from rat urine, namely, 5-hydroxy-pulegone (VIII), piperitone (IX), piperitenone (X), 7-hydroxy-piperitone (XI), 8-hydroxy piperitone (XII), p-cresol (XIII), geranic acid (XIV) and neronic acid (XV). These were identified by n.m.r., i.r. and mass spectrometry. 3. Based on these results, metabolic pathways for the biotransformation of R-(+)-pulegone in rat have been proposed.

Effect of myonuclear number and mitochondrial fusion on Drosophila indirect flight muscle organization and size

Mechanisms involved in establishing the organization and numbers of fibres in a muscle are not completely understood. During Drosophila indirect flight muscle (IFM) formation, muscle growth is achieved by both incorporating hundreds of nuclei, and hypertrophy. As a result, IFMs provide a good model with which to understand the mechanisms that govern overall muscle organization and growth. We present a detailed analysis of the organization of dorsal longitudinal muscles (DLMs), a subset of the IFMs. We show that each DLM is similar to a vertebrate fascicle and consists of multiple muscle fibres. However, increased fascicle size does not necessarily change the number of constituent fibres, but does increase the number of myofibrils packed within the fibres. We also find that altering the number of myoblasts available for fusion changes DLM fascicle size and fibres are loosely packed with myofibrils. Additionally, we show that knock down of genes required for mitochondrial fusion causes a severe reduction in the size of DLM fascicles and fibres. Our results establish the organization levels of DLMs and highlight the importance of the appropriate number of nuclei and mitochondrial fusion in determining the overall organization, growth and size of DLMs. (C) 2013 Elsevier Inc. All rights reserved.

Single-ion magnet behaviour of heptacoordinated Fe(II) complexes: on the importance of supramolecular organization

Supramolecular organization of a metal complex may significantly contribute to the magnetization dynamics of mononuclear SMMs. This is illustrated for a heptacoordinated Fe(II) complex with rather moderate Ising-type anisotropy for which a slow magnetization relaxation with significant energy barrier was reached when this complex was properly organized in the crystal lattice. Incidentally, it is the first example of single-ion magnet behaviour of Fe(II) in a pentagonal bipyramid surrounding.

User’s Guide to ADMB2R: A Set of AD Model Builder Output Routines Compatible with the R Statistics Language

ADMB2R is a collection of AD Model Builder routines for saving complex data structures into a file that can be read in the R statistics environment with a single command.1 ADMB2R provides both the means to transfer data structures significantly more complex than simple tables, and an archive mechanism to store data for future reference. We developed this software because we write and run computationally intensive numerical models in Fortran, C++, and AD Model Builder. We then analyse results with R. We desired to automate data transfer to speed diagnostics during working-group meetings. We thus developed the ADMB2R interface to write an R data object (of type list) to a plain-text file. The master list can contain any number of matrices, values, dataframes, vectors or lists, all of which can be read into R with a single call to the dget function. This allows easy transfer of structured data from compiled models to R. Having the capacity to transfer model data, metadata, and results has sharply reduced the time spent on diagnostics, and at the same time, our diagnostic capabilities have improved tremendously. The simplicity of this interface and the capabilities of R have enabled us to automate graph and table creation for formal reports. Finally, the persistent storage in files makes it easier to treat model results in analyses or meta-analyses devised months—or even years—later. We offer ADMB2R to others in the hope that they will find it useful. (PDF contains 30 pages)

User’s Guide to C2R: A Set of C Language Output Routines Compatible with the R Statistics Language

C2R is a collection of C routines for saving complex data structures into a file that can be read in the R statistics environment with a single command.1 C2R provides both the means to transfer data structures significantly more complex than simple tables, and an archive mechanism to store data for future reference. We developed this software because we write and run computationally intensive numerical models in Fortran, C++, and AD Model Builder. We then analyse results with R. We desired to automate data transfer to speed diagnostics during working-group meetings. We thus developed the C2R interface to write an R data object (of type list) to a plain-text file. The master list can contain any number of matrices, values, dataframes, vectors or lists, all of which can be read into R with a single call to the dget function. This allows easy transfer of structured data from compiled models to R. Having the capacity to transfer model data, metadata, and results has sharply reduced the time spent on diagnostics, and at the same time, our diagnostic capabilities have improved tremendously. The simplicity of this interface and the capabilities of R have enabled us to automate graph and table creation for formal reports. Finally, the persistent storage in files makes it easier to treat model results in analyses or meta-analyses devised months—or even years—later. We offer C2R to others in the hope that they will find it useful. (PDF contains 27 pages)

User’s Guide to For2R: A Module of Fortran 95 Output Routines Compatible with the R Statistics Language

For2R is a collection of Fortran routines for saving complex data structures into a file that can be read in the R statistics environment with a single command.1 For2R provides both the means to transfer data structures significantly more complex than simple tables, and an archive mechanism to store data for future reference. We developed this software because we write and run computationally intensive numerical models in Fortran, C++, and AD Model Builder. We then analyse results with R. We desired to automate data transfer to speed diagnostics during working-group meetings. We thus developed the For2R interface to write an R data object (of type list) to a plain-text file. The master list can contain any number of matrices, values, dataframes, vectors or lists, all of which can be read into R with a single call to the dget function. This allows easy transfer of structured data from compiled models to R. Having the capacity to transfer model data, metadata, and results has sharply reduced the time spent on diagnostics, and at the same time, our diagnostic capabilities have improved tremendously. The simplicity of this interface and the capabilities of R have enabled us to automate graph and table creation for formal reports. Finally, the persistent storage in files makes it easier to treat model results in analyses or meta-analyses devised months—or even years—later. We offer For2R to others in the hope that they will find it useful. (PDF contains 31 pages)

The fishing centres of Lake Niassa (Mozambique): results of a frame survey made in June 1983

Results of the first frame survey of the fishing centres on the Mozambican coast of Lake Niassa, held in June 1983, are presented. A total of 41 fishing centres were detected in the area and visited. Through direct enquiries in each centre, an estimated number of 3,380 fishermen and 1,230 fishing boats (of which 25 motorized) was established. The results also provided information on the types and quantities of fishing gear used. A first estimation of the approximate total annual catch in the area gave a figure in the order of 9,100 tons., of which approximately equals 90% is landed during the rainy season. The survey is a part of a research programme on the fishing resources of Lake Niassa and of a project for the collection and organization of fisheries statistics from the artisanal fisheries of the Mozambican side of Lake Niassa.

Characterization and Comparison of the Tissue-Related Modules in Human and Mouse

Background: Due to the advances of high throughput technology and data-collection approaches, we are now in an unprecedented position to understand the evolution of organisms. Great efforts have characterized many individual genes responsible for the interspecies divergence, yet little is known about the genome-wide divergence at a higher level. Modules, serving as the building blocks and operational units of biological systems, provide more information than individual genes. Hence, the comparative analysis between species at the module level would shed more light on the mechanisms underlying the evolution of organisms than the traditional comparative genomics approaches. Results: We systematically identified the tissue-related modules using the iterative signature algorithm (ISA), and we detected 52 and 65 modules in the human and mouse genomes, respectively. The gene expression patterns indicate that all of these predicted modules have a high possibility of serving as real biological modules. In addition, we defined a novel quantity, "total constraint intensity,'' a proxy of multiple constraints (of co-regulated genes and tissues where the co-regulation occurs) on the evolution of genes in module context. We demonstrate that the evolutionary rate of a gene is negatively correlated with its total constraint intensity. Furthermore, there are modules coding the same essential biological processes, while their gene contents have diverged extensively between human and mouse. Conclusions: Our results suggest that unlike the composition of module, which exhibits a great difference between human and mouse, the functional organization of the corresponding modules may evolve in a more conservative manner. Most importantly, our findings imply that similar biological processes can be carried out by different sets of genes from human and mouse, therefore, the functional data of individual genes from mouse may not apply to human in certain occasions.